A T7 RNA polymerase-dependent gene expression system for Bacillus megaterium
| Autor: | David Fröde, Rebekka Biedendieck, Martin Gamer, Simon Stammen, Dieter Jahn |
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| Rok vydání: | 2009 |
| Předmět: |
Models
Molecular Genetic Vectors Green Fluorescent Proteins Molecular Sequence Data Gene Expression Biology Applied Microbiology and Biotechnology Green fluorescent protein chemistry.chemical_compound Viral Proteins Bacterial Proteins RNA polymerase Bacteriophage T7 Gene expression medicine T7 RNA polymerase Bacillus megaterium Base Sequence fungi RNA Levansucrase General Medicine DNA-Directed RNA Polymerases biology.organism_classification Molecular biology Recombinant Proteins Secretory protein chemistry Hexosyltransferases Nucleic Acid Conformation Biotechnology medicine.drug |
| Zdroj: | Applied microbiology and biotechnology. 82(6) |
| ISSN: | 1432-0614 |
| Popis: | Gene expression systems based on the RNA polymerase of the bacteriophage T7 are often the ultimate choice for the high level production of recombinant proteins. During the last decade, the Gram-positive bacterium Bacillus megaterium was established as a useful host for the intra- and extracellular production of heterologous proteins. In this paper, we report on the development of a T7 RNA polymerase-dependent expression system for B. megaterium. The system was evaluated for cytosolic and secretory protein production with green fluorescent protein (GFP) from Aequoria victoria as intracellular and Lactobacillus reuteri levansucrase as extracellular model protein. GFP accumulated rapidly at high levels up to 50 mg/l shake flask culture intracellularly after induction of T7 RNA polymerase gene expression. The addition of rifampicin for the inhibition of B. megaterium RNA polymerase led to an increased stability of GFP. L. reuteri levansucrase was also successfully produced and secreted (up to 20 U/l) into the culture supernatant. However, parallel intracellular accumulation of the protein indicated limitations affiliated with the Sec-dependent protein translocation process. |
| Databáze: | OpenAIRE |
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