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Media type:
E-Article
Title:
Improved Plasmid-Based Inducible and Constitutive Gene Expression in Corynebacterium glutamicum
Contributor:
Henke, Nadja A.;
Krahn, Irene;
Wendisch, Volker F.
Published:
MDPI AG, 2021
Published in:
Microorganisms, 9 (2021) 1, Seite 204
Language:
English
DOI:
10.3390/microorganisms9010204
ISSN:
2076-2607
Origination:
Footnote:
Description:
Corynebacterium glutamicum has been safely used in white biotechnology for the last 60 years and the portfolio of new pathways and products is increasing rapidly. Hence, expression vectors play a central role in discovering endogenous gene functions and in establishing heterologous gene expression. In this work, new expression vectors were designed based on two strategies: (i) a library screening of constitutive native and synthetic promoters and (ii) an increase of the plasmid copy number. Both strategies were combined and resulted in a very strong expression and overproduction of the fluorescence protein GfpUV. As a second test case, the improved vector for constitutive expression was used to overexpress the endogenous xylulokinase gene xylB in a synthetic operon with xylose isomerase gene xylA from Xanthomonas campestris. The xylose isomerase activity in crude extracts was increased by about three-fold as compared to that of the parental vector. In terms of application, the improved vector for constitutive xylA and xylB expression was used for production of the N-methylated amino acid sarcosine from monomethylamine, acetate, and xylose. As a consequence, the volumetric productivity of sarcosine production was 50% higher as compared to that of the strain carrying the parental vector.