In order to improve the 4-hydroxyisoleucine (4-HIL) fermentation ability of Corynebacterium glutamicum and to reduce the use of antibiotics, the alanine racemase encoding gene alr was knocked out in C. glutamicum strain SN01, and the plasmid pJYW4-ido expressing alr gene and ido gene was supplemented, yielding the strain Δalr/p4-ido. Experiment results showed that this method could bring enough selecting pressure to cells, and the plasmids could be maintained to produce 4-HIL without the use of antibiotics during fermentation. Then the plasmid pJYW4-ido-lysC-ido3 was transformed into SN01Δalr, yielding strain Δalr-ILI3 which can be fermented without antibiotics. The fed-batch fermentation conditions of this strain were optimized. Firstly, seeds with different growth time were inoculated for fed-batch fermentation. The results showed that seeds with a growth time of 18 h were most suitable for 4-HIL production. After 96 h of fermentation, 242.18 mmol/L 4-HIL was produced. Secondly, the dissolved oxygen level was adjusted in two stages, and the 4-HIL production reached 269.20 mmol/L after 96 h fermentation. The titer increased by 11.2% compared with that before optimization. Finally, after adjusting the feeding method, the production of 4-HIL reached 324.72 mmol/L, which was 34.1% higher than that before optimization. This method provided a strategy for performing 4-HIL fermentation without antibiotics and for improving 4-HIL production in C. glutamicum.
WEI Shuyu
,
SHI Feng
. Construction of strains for 4-hydroxyisoleucine fermentation without antibiotics and optimization of fermentation conditions[J]. Food and Fermentation Industries, 2022
, 48(12)
: 117
-122
.
DOI: 10.13995/j.cnki.11-1802/ts.030983
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