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Development of a plasmid-based, tunable, tolC-derived expression system for application in Cupriavidus necator H16

  • Cupriavidus necator H16 gains increasing attention in microbial research and biotechnological application due to its diverse metabolic features. Here we present a tightly controlled gene expression system for C. necator including the pBBR1-vector that contains hybrid promoters originating from C. necator native tolC-promoter in combination with a synthetic tetO-operator. The expression of the reporter gene from these plasmids relies on the addition of the exogenous inducer doxycycline (dc). The novel expression system offers a combination of advantageous features as; (i) high and dose-dependent recombinant protein production, (ii) tight control with a high dynamic range (On/Off ratio), which makes it applicable for harmful pathways or for toxic protein production, (iii) comparable cheap inducer (doxycycline, dc), (iv) effective at low inducer concentration, that makes it useful for large scale application, (v) rapid, diffusion controlled induction, and (vi) the inducer does not interfere within the cell metabolism. As applications of the expression system in C. necator H16, the growth ability on glycerol was enhanced by constitutively expressing the E. coli glpk gene-encoding for glycerol kinase. Likewise, we used the system to overcome the expression toxicity of mevalonate pathway in C. necator H16. With this system, the mevalonate-genes were successfully introduced in the host and the recombinant strains could produce about 200 mg/l mevalonate.

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Metadaten
Author:E. A. Aboulnaga, H. Zou, Thorsten Selmer, M. Xian
DOI:https://doi.org/10.1016/j.jbiotec.2018.03.007
ISSN:0168-1656
Parent Title (English):Journal of Biotechnology
Publisher:Elsevier
Place of publication:Amsterdam
Document Type:Article
Language:English
Year of Completion:2018
Date of the Publication (Server):2018/04/30
Volume:274
First Page:15
Last Page:27
Link:http://doi.org/10.1016/j.jbiotec.2018.03.007
Zugriffsart:bezahl
Institutes:FH Aachen / Fachbereich Chemie und Biotechnologie
FH Aachen / INB - Institut für Nano- und Biotechnologien
collections:Verlag / Elsevier