Abstract
BackgroundBetaine, an osmoprotective compatible solute, has been used to improve l-threonine production in engineered Escherichia colil-threonine producer. Betaine supplementation upregulates the expression of zwf encoding glucose-6-phosphate dehydrogenase, leading to the increase of NADPH, which is beneficial for l-threonine production. In E. coli, betaine can be taken through ProP encoded by proP or ProVWX encoded by proVWX. ProP is a H+-osmolyte symporter, whereas ProVWX is an ABC transporter. ProP and ProVWX mediate osmotic stress protection by transporting zwitterionic osmolytes, including glycine betaine. Betaine can also be synthesized in E. coli by enzymes encoded by betABIT. However, the influence of ProP, ProVWX and betABIT on l-threonine production in E. coli has not been investigated.ResultsIn this study, the influence of ProP, ProVWX and betABIT on l-threonine production in E. coli has been investigated. Addition of betaine slightly improved the growth of the l-threonine producing E. coli strain TWF001 as well as the l-threonine production. Deletion of betABIT retarded the growth of TWF001 and slightly decreased the l-threonine production. However, deletion of proP or/and proVWX significantly increased the l-threonine production. When proP was deleted, the l-threonine production increased 33.3%; when proVWX was deleted, the l-threonine production increased 40.0%. When both proP and proVWX were deleted, the resulting strain TSW003 produced 23.5 g/l l-threonine after 36 h flask cultivation. The genes betABIT, proC, fadR, crr and ptsG were individually deleted from TSW003, and it was found that further absence of either crr (TWS008) or ptsG (TWS009) improved l-threonine production. TSW008 produced 24.9 g/l l-threonine after 36 h flask cultivation with a yield of 0.62 g/g glucose and a productivity of 0.69 g/l/h. TSW009 produced 26 g/l l-threonine after 48 h flask cultivation with a yield of 0.65 g/g glucose and a productivity of 0.54 g/l/h, which is 116% increase compared to the control TWF001.ConclusionsIn this study, l-threonine-producing E. coli strains TSW008 and TSW009 with high l-threonine productivity were developed by regulating the intracellular osmotic pressure. This strategy could be used to improve the production of other products in microorganisms.
Highlights
In the process of fermentation production, we often encounter the growth restriction and the termination of product biosynthesis due to the osmotic pressure imbalance
To investigate the influence of betaine on l-threonine production of TWF001, 0.5, 1.0, 1.5 and 2.0 g/l betaine were added during fermentation [4], the cell growth, glucose consumption, and l-threonine production were determined and compared (Fig. 2)
The results suggest that the addition of betaine can influence the production of l-threonine in TWF001 possibly due to the change of osmotic pressure
Summary
In the process of fermentation production, we often encounter the growth restriction and the termination of product biosynthesis due to the osmotic pressure imbalance. Improving the ability of bacteria to tolerate osmotic stress or using special strains that. The resulting recombinant strain produced 33 g/l l-threonine when grown under open unsterile conditions with a productivity of 1.368 g/l/h in fed-batch fermentation [2]. Osmoprotectants have been used to improve the growth and yield limitations caused by the imbalance of osmotic pressure during the fermentation process [3, 4]. Since hypertonic tolerant strains need complex screening and genetic modification, modification of the existing high-yielding bacteria to improve their osmotic resistance is a good strategy [8, 9]. The influence of ProP, ProVWX and betABIT on l-threonine production in E. coli has not been investigated
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