Abstract
BackgroundL-ornithine is effective in the treatment of liver diseases and helps strengthen the heart. The commercial applications mean that efficient biotechnological production of L-ornithine has become increasingly necessary. Adaptive evolution strategies have been proven a feasible and efficient technique to achieve improved cellular properties without requiring metabolic or regulatory details of the strain. The evolved strains can be further optimised by metabolic engineering. Thus, metabolic evolution strategy was used for engineering Corynebacterium glutamicum to enhance L-ornithine production.ResultsA C. glutamicum strain was engineered by using a combination of gene deletions and adaptive evolution with 70 passages of growth-based selection. The metabolically evolved C. glutamicum strain, named ΔAPE6937R42, produced 24.1 g/L of L-ornithine in a 5-L bioreactor. The mechanism used by C. glutamicum ΔAPE6937R42 to produce L-ornithine was investigated by analysing transcriptional levels of select genes and NADPH contents. The upregulation of the transcription levels of genes involved in the upstream pathway of glutamate biosynthesis and the elevated NADPH concentration caused by the upregulation of the transcriptional level of the ppnK gene promoted L-ornithine production in C. glutamicum ΔAPE6937R42.ConclusionsThe availability of NADPH plays an important role in L-ornithine production in C. glutamicum. Our results demonstrated that the combination of growth-coupled evolution with analysis of transcript abundances provides a strategy to engineer microbial strains for improving production of target compounds.
Highlights
L-ornithine is effective in the treatment of liver diseases and helps strengthen the heart
Lee and Cho reported that an engineered Escherichia coli produced 13.2 mg L-ornithine per gram of dry cell weight (DCW), and that addition of glutamate to the culture favoured L-ornithine production in the engineered E. coli [8]
Hwang et al reported that cooverexpression of argCJBD in a triple-gene knockout strain C. glutamicum ATCC 13032 (ΔargFΔargRΔproB) resulted in a cellular L-ornithine content of 16.49 mg/g DCW and a concentration of L-ornithine in the culture medium of 179.14 mg/L [9]
Summary
L-ornithine is effective in the treatment of liver diseases and helps strengthen the heart. L-ornithine, a non-essential amino acid and an important constituent of the urea cycle, is the precursor of other amino acids, such as citrulline and arginine It is effective for the treatment and prophylaxis of liver diseases [1], and has been applied to wound healing [2]. The L-ornithine producing strain C. glutamicum ATCC 13032 (ΔargFΔargR) named ORN1 was constructed and shown to produce L-ornithine from arabinose when araBAD from E. coli was expressed [13]. This group constructed an engineered C. glutamicum ORN1 (pEKEx3-xylAXc-xylBCg) to effectively produce Lornithine from xylose [14]. This strain, named ATCC13032 (ΔargFΔproBΔkgd), produced L-ornithine of 18.17 g/L in the optimal medium [16]
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