High-cell-density fermentation for production ofL-N-carbamoylase using an expression system based on theEscherichia coli rhaBAD promoter
Autor: | Matthias Reuss, Burkhard Wilms, Martin Siemann, Ralf Mattes, Josef Altenbuchner, Achim Hauck, Christoph Syldatk |
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Rok vydání: | 2001 |
Předmět: |
Rhamnose
Cell Count Bioengineering medicine.disease_cause Applied Microbiology and Biotechnology Amidohydrolases chemistry.chemical_compound Plasmid Escherichia coli medicine Inducer Arthrobacter Promoter Regions Genetic Amino Acid Isomerases ColE1 Expression vector biology Gene Expression Regulation Bacterial biology.organism_classification Enterobacteriaceae Biochemistry chemistry Fermentation Plasmids Biotechnology |
Zdroj: | Biotechnology and Bioengineering. 73:95-103 |
ISSN: | 1097-0290 0006-3592 |
Popis: | A high-cell-density fed-batch fermentation for the production of heterologous proteins in Escherichia coli was developed using the positively regulated Escherichia coli rhaBAD promoter. The expression system was improved by reducing of the amount of expensive L-rhamnose necessary for induction of the rhamnose promoter and by increasing the vector stability. Consumption of the inducer L-rhamnose was inhibited by inactivation of L-rhamnulose kinase encoding gene rhaB of Escherichia coli W3110, responsible for the first irreversible step in rhamnose catabolism. Plasmid instability caused by multimerization of the expression vector in the recombination-proficient W3110 was prevented by insertion of the multimer resolution site cer from the ColE1 plasmid into the vector. Fermentation experiments with the optimized system resulted in the production of 100 g x L(-1) cell dry weight and 3.8 g x L(-1) of recombinant L-N-carbamoylase, an enzyme, which is needed for the production of enantiomeric pure amino acids in a two-step reaction from hydantoins. |
Databáze: | OpenAIRE |
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