Biotechnology and Bioengineering (BB) | Enzyme, Protein Engineering, and Metabolic Engineering
Effect of the pat, fk, stpk gene knock-out and mdh gene knock-in on mannitol production in Leuconostoc mesenteroides
School of Chemical Engineering and Technology, Hebei University of Technology
Received: May 28, 2018; Accepted: October 5, 2018
J. Microbiol. Biotechnol. 2018; 28(12): 2009-2018
Published December 28, 2018
Copyright © The Korean Society for Microbiology and Biotechnology.
Leuconostoc mesenteroides can be used to produce mannitol by fermentation, but the mannitol productivity is not high. Therefore, this study modify the chromosome of Leuconostoc mesenteroides by genetic methods to obtain high-yield strains of mannitol. In this study, gene knock-out strains and gene knock-in strains were constructed by a two-step homologous recombination method. The mannitol productivity of the pat gene (which encoding phosphate acetyltransferase) deleteon strain (Δpat::amy), fk gene (which encoding fructokinase) deleteon strain (Δfk::amy) and stpk gene (which encoding serine-threonine protein kinase) deleteon strain (Δstpk::amy) were all increased compared to the wild type, and the productivity of mannitol were 84.8 %, 83.5 % and 84.1% respectively. The mannitol productivity of mdh gene (which encoding mannitol dehydrogenase) knock-in strain Δpat::mdh, Δfk::mdh and Δstpk::mdh was increased than the single gene deletion strains, and the productivity of mannitol was 96.5 %, 88 % and 93.2 % respectively. The multi-mutant strain ΔdtsΔldhΔpat::mdhΔstpk::mdhΔfk::mdh with a mannitol productivity of 97.3 %. This work shows that multi-gene knock-out and gene knock-in strains have the greatest impact on mannitol production, with a mannitol productivity of 97.3% and an increase of 24.7 % over wild type. This study used the methods of gene knock-out and gene knock-in to genetically modify the chromosome of Leuconostoc mesenteroides. It is of great significance to increase the synthesis ability of mannitol and has broad development prospects.
Leuconostoc mesenteroides, mannitol Productivity, gene knock-out, gene knock-in
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