Effect of postinduction nutrient feed composition and use of lactose as inducer during production of thermostable xylanase in Escherichia coli glucose-limited fed-batch cultivations

J Biosci Bioeng. 2005 May;99(5):477-84. doi: 10.1263/jbb.99.477.

Abstract

Escherichia coli is a microorganism routinely used in the production of heterologous proteins. The overexpression of a xylanase (Xyn 10 A Delta NC), which originated from the thermophile Rhodothermus marinus cloned under the control of the strong T7/lac promoter in a defined medium (mAT) using a substrate-limited feed strategy, was however shown to impose a significant metabolic burden on host cells. This resulted in a decreased cell growth rate and ultimately also a decreased target protein production. The investigation hence centers on the effect of some selected nutrient feed additives (amino acid [Cys] or TCA-intermediates [citrate, succinate, malate]) used to relieve the metabolic burden imposed during the feeding and postinduction phases of these glucose-limited fed-batch cultivations. The use of either succinic acid or malic acid as feed-additives resulted in an increase in production of approximately 40% of the heterologous thermostable xylanase. Furthermore, use of lactose as an alternative inducer of the T7/lac promoter was also proven to be a suitable strategy that significantly prolonged the heterologous protein production phase as compared with induction using isopropyl beta-D-thiogalactopyranoside (IPTG).

Publication types

  • Comparative Study
  • Evaluation Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bioreactors / microbiology*
  • Cell Culture Techniques / methods*
  • Endo-1,4-beta Xylanases / biosynthesis*
  • Endo-1,4-beta Xylanases / genetics
  • Enzyme Activation
  • Enzyme Stability
  • Escherichia coli / enzymology*
  • Escherichia coli / genetics
  • Glucose / metabolism*
  • Lactose / metabolism*
  • Protein Engineering / methods*
  • Recombinant Proteins / metabolism
  • Temperature

Substances

  • Recombinant Proteins
  • Endo-1,4-beta Xylanases
  • Glucose
  • Lactose