Genetically switched D-lactate production in Escherichia coli.
During a fermentation process, the formation of the desired product during the cell growth phase competes with the biomass for substrates or inhibits cell growth directly, which results in a decrease in production efficiency. A genetic switch is required to precisely separate growth from production and to simplify the fermentation process. The ldhA promoter, which encodes the fermentative D-lactate dehydrogenase (LDH) in the lactate producer Escherichia coli CICIM B0013-070 (ack-pta pps pflB dld poxB adhE frdA), was replaced with the λ p(R) and p(L) promoters (as a genetic switch) using genomic recombination and the thermo-controllable strain B0013-070B (B0013-070, ldhAp::kan-cI(ts)857-p(R)-p(L)), which could produce two-fold higher LDH activity at 42°C than the B0013-070 strain, was created. When the genetic switch was turned off at 33°C, strain B0013-070B produced 10% more biomass aerobically than strain B0013-070 and produced only trace levels of lactate which could reduce the growth inhibition caused by oxygen insufficiency in large scale fermentation. However, 42°C is the most efficient temperature for switching on lactate production. The volumetric productivity of B0013-070B improved by 9% compared to that of strain B0013-070 when it was grown aerobically at 33°C with a short thermo-induction at 42°C and then switched to the production phase at 42°C. In a bioreactor experiment using scaled-up conditions that were optimized in a shake flask experiment, strain B0013-070B produced 122.8 g/l D-lactate with an increased oxygen-limited productivity of 0.89 g/g·h. The results revealed the effectiveness of using a genetic switch to regulate cell growth and the production of a metabolic compound.
Center for Bioresource & Bioenergy, School of Biotechnology, Jiangnan University, 1800 Lihu Avenue, Wuxi 214122, People's Republic of China.
Li
L
Zhou
Dan-Dan
DD
Niu
Kang-Ming
KM
Tian
Xian-Zhong
XZ
Chen
Bernard A
BA
Prior
Wei
W
Shen
Gui-Yang
GY
Shi
Suren
S
Singh
Zheng-Xiang
ZX
Wang
D000332:Aerobiosis / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D029968:Escherichia coli Proteins / Q000096:biosynthesis / Q000235:genetics; D005818:Genetic Engineering; D007770:L-Lactate Dehydrogenase / Q000096:biosynthesis / Q000235:genetics; D019344:Lactic Acid / Q000096:biosynthesis*; D011401:Promoter Regions, Genetic / Q000235:genetics
D029968:Escherichia coli Proteins; D019344:Lactic Acid; D007770:L-Lactate Dehydrogenase