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Recruiting alternative glucose utilization pathways for improving succinate production.

Jinlei Tang1, Xinna Zhu, Jiao Lu

  • 1Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.

Applied Microbiology and Biotechnology
|August 17, 2012
PubMed
Summary
This summary is machine-generated.

Inactivating the phosphoenolpyruvate (PEP): carbohydrate phosphotransferase system (PTS) in E. coli hinders growth. This study engineered PTS-negative strains with alternative glucose transporters (Glf or GalP) and glucokinases (Glk) to enhance glucose utilization and succinate production.

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Area of Science:

  • Metabolic Engineering
  • Synthetic Biology
  • Biotechnology

Background:

  • The phosphoenolpyruvate (PEP): carbohydrate phosphotransferase system (PTS) is crucial for glucose uptake in Escherichia coli.
  • PTS inactivation is a strategy to increase PEP availability for succinate production, but it impairs cell growth and glucose metabolism.
  • Alternative glucose transport and phosphorylation pathways are needed to overcome limitations in PTS-deficient strains.

Purpose of the Study:

  • To engineer and compare alternative glucose utilization pathways in PTS-negative E. coli strains.
  • To evaluate the impact of different glucose transporters (Z. mobilis Glf vs. E. coli GalP) and glucokinases (E. coli Glk vs. Z. mobilis Glk) on glucose consumption and succinate production.
  • To identify optimal combinations for enhanced microbial production of PEP-derived chemicals.

Main Methods:

  • Construction of PTS-negative E. coli strains.
  • Heterologous expression of glucose facilitators (Glf) and permeases (GalP).
  • Co-expression of different glucokinases (Glk) from E. coli and Zymomonas mobilis.
  • Measurement of glucose utilization rates and succinate production yields.
  • Comparative analysis of engineered strains against wild-type E. coli.

Main Results:

  • PTS-negative strains expressing Z. mobilis Glf exhibited significantly higher glucose utilization rates compared to those with E. coli GalP.
  • Combinatorial modulation of Glf and E. coli Glk resulted in an 81% increase in glucose utilization rate over wild-type E. coli.
  • Succinate productivity increased by 20% with GalP and 41% with Glf.
  • Replacing E. coli Glk with Z. mobilis Glk, despite higher activity, led to decreased glucose utilization, likely due to energy-dependent regulation.

Conclusions:

  • Z. mobilis Glf is a more efficient glucose transporter for PTS-negative E. coli than E. coli GalP.
  • Engineered alternative glucose pathways significantly enhance glucose utilization and succinate production.
  • These strategies provide a robust platform for the biotechnological production of various PEP-derived chemicals.