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Enhanced Exopolysaccharide Production by Metabolic Engineering of Streptococcus thermophilus.

Levander, Fredrik LU ; Svensson, Malin LU and Rådström, Peter LU (2002) In Applied and Environmental Microbiology 68(2). p.784-790
Abstract
It is possible that the low levels of production of exopolysaccharides (EPSs) by lactic acid bacteria could be improved by altering the levels of enzymes in the central metabolism that influence the production of precursor nucleotide sugars. To test this hypothesis, we identified and cloned the galU gene, which codes for UDP glucose pyrophosphorylase (GalU) in Streptococcus thermophilus LY03. Homologous overexpression of the gene led to a 10-fold increase in GalU activity but did not have any effect on the EPS yield when lactose was the carbon source. However, when galU was overexpressed in combination with pgmA, which encodes phosphoglucomutase (PGM), the EPS yield increased from 0.17 to 0.31 g/mol of carbon from lactose. A... (More)
It is possible that the low levels of production of exopolysaccharides (EPSs) by lactic acid bacteria could be improved by altering the levels of enzymes in the central metabolism that influence the production of precursor nucleotide sugars. To test this hypothesis, we identified and cloned the galU gene, which codes for UDP glucose pyrophosphorylase (GalU) in Streptococcus thermophilus LY03. Homologous overexpression of the gene led to a 10-fold increase in GalU activity but did not have any effect on the EPS yield when lactose was the carbon source. However, when galU was overexpressed in combination with pgmA, which encodes phosphoglucomutase (PGM), the EPS yield increased from 0.17 to 0.31 g/mol of carbon from lactose. A galactose-fermenting LY03 mutant (Gal(+)) with increased activities of the Leloir enzymes was also found to have a higher EPS yield (0.24 g/mol of carbon) than the parent strain. The EPS yield was further improved to 0.27 g/mol of carbon by overexpressing galU in this strain. However, the highest EPS yield, 0.36 g/mol of carbon, was obtained when pgmA was knocked out in the Gal(+) strain. Measurements of the levels of intracellular metabolites in the cultures revealed that the Gal(+) strains had considerably higher glucose 1-phosphate levels than the other strains, and the strain lacking PGM activity had threefold-higher levels of glucose 1-phosphate than the other Gal(+) strains. These results show that it is possible to increase EPS production by altering the levels of enzymes in the central carbohydrate metabolism. (Less)
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organization
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Contribution to journal
publication status
published
subject
keywords
Genetic Engineering/*methods, Molecular Sequence Data, Phosphoglucomutase/genetics/metabolism, UTP-Glucose-1-Phosphate Uridylyltransferase/*genetics/*metabolism, Non-U.S. Gov't, Support, Streptococcus/*enzymology/genetics/growth & development, Polysaccharides, Bacterial/*biosynthesis, Galactose/metabolism, Molecular, Culture Media, Cloning, Base Sequence
in
Applied and Environmental Microbiology
volume
68
issue
2
pages
784 - 790
publisher
American Society for Microbiology
external identifiers
  • wos:000173588600043
  • pmid:11823219
  • scopus:0036156924
ISSN
0099-2240
DOI
10.1128/AEM.68.2.784-790.2002
language
English
LU publication?
yes
id
6f438808-8cdd-47b7-ad1e-656bd03220b4 (old id 106238)
date added to LUP
2007-06-28 15:54:53
date last changed
2017-10-08 03:40:41
@article{6f438808-8cdd-47b7-ad1e-656bd03220b4,
  abstract     = {It is possible that the low levels of production of exopolysaccharides (EPSs) by lactic acid bacteria could be improved by altering the levels of enzymes in the central metabolism that influence the production of precursor nucleotide sugars. To test this hypothesis, we identified and cloned the galU gene, which codes for UDP glucose pyrophosphorylase (GalU) in Streptococcus thermophilus LY03. Homologous overexpression of the gene led to a 10-fold increase in GalU activity but did not have any effect on the EPS yield when lactose was the carbon source. However, when galU was overexpressed in combination with pgmA, which encodes phosphoglucomutase (PGM), the EPS yield increased from 0.17 to 0.31 g/mol of carbon from lactose. A galactose-fermenting LY03 mutant (Gal(+)) with increased activities of the Leloir enzymes was also found to have a higher EPS yield (0.24 g/mol of carbon) than the parent strain. The EPS yield was further improved to 0.27 g/mol of carbon by overexpressing galU in this strain. However, the highest EPS yield, 0.36 g/mol of carbon, was obtained when pgmA was knocked out in the Gal(+) strain. Measurements of the levels of intracellular metabolites in the cultures revealed that the Gal(+) strains had considerably higher glucose 1-phosphate levels than the other strains, and the strain lacking PGM activity had threefold-higher levels of glucose 1-phosphate than the other Gal(+) strains. These results show that it is possible to increase EPS production by altering the levels of enzymes in the central carbohydrate metabolism.},
  author       = {Levander, Fredrik and Svensson, Malin and Rådström, Peter},
  issn         = {0099-2240},
  keyword      = {Genetic Engineering/*methods,Molecular Sequence Data,Phosphoglucomutase/genetics/metabolism,UTP-Glucose-1-Phosphate Uridylyltransferase/*genetics/*metabolism,Non-U.S. Gov't,Support,Streptococcus/*enzymology/genetics/growth & development,Polysaccharides,Bacterial/*biosynthesis,Galactose/metabolism,Molecular,Culture Media,Cloning,Base Sequence},
  language     = {eng},
  number       = {2},
  pages        = {784--790},
  publisher    = {American Society for Microbiology},
  series       = {Applied and Environmental Microbiology},
  title        = {Enhanced Exopolysaccharide Production by Metabolic Engineering of Streptococcus thermophilus.},
  url          = {http://dx.doi.org/10.1128/AEM.68.2.784-790.2002},
  volume       = {68},
  year         = {2002},
}