Optimization of fed-batch fermentation for a staphylokinase-hirudin fusion protein in Escherichia coli BL21

  • Zhang G
  • Zhong G
  • Wang X
 et al. 
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Abstract

In this study, the fed-batch fermentation technique was applied to improve the yield of STH, a chimeric protein composed of staphylokinase (SAK) and hirudin (HV) with thrombin recognition peptide as a linker peptide, produced by Escherichia coli BL21. Three kinds of feeding mediums, GM (glucose + magnesium sulfate), GMY (GM + yeast extract) and GMYT (GMY + tryptone), were investigated with R-medium as an initial culture medium in a 5 L fermentor. The results showed that the dry cell weight were similar in the three feeding mediums, but the yield of total protein and STH were significantly different (total protein: GM, GMY, GMYT = 1, 4.72, 7.36; STH: GM, GMY, GMYT = 1, 1.93, 3.23). Then we replace the R-medium with the complex medium which contains yeast extract and tryptone in fed-batch fermentation based on the GMYT as feeding medium. The results showed that the total protein and STH in the complex medium were 6.29 and 7.76 fold of those in R-medium culturing condition, respectively. Under optimal conditions (GMYT and complex medium), a final STH expression of 1.48 g/l fermentation broth was achieved after 20 h cultivation. The efficacy of STH production was 37 times more than that before technique optimization. Furthermore, the optimal conditions were successfully scaled up to 40 L fermentor. This may prompt STH to develop into a new thrombolytic agent. It is interesting to know that for the harvest, the yeast extract and tryptone can significantly contribute to the protein expression.

Author-supplied keywords

  • ANTITHROMBOTIC AGENT
  • CULTIVATION
  • Complex medium
  • GROWTH-RATE
  • HIGH CELL-DENSITY
  • HIGHER FIBRIN AFFINITY
  • PLATSAK
  • R-medium
  • VARIANT
  • fed-batch fermentation
  • feeding medium

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Authors

  • Guangman Zhang

  • Genshen Zhong

  • Xiudong Wang

  • Lisheng Wang

  • Yide Qin

  • Aiping Yu

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