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Rational design and metabolic analysis of escherichia coli for effective production of L-tryptophan at high concentration
Publikationstyp
Journal Article
Date Issued
2016-09-06
Sprache
English
Author(s)
Institut
TORE-URI
Volume
101
Issue
2
Start Page
559
End Page
568
Citation
Applied Microbiology and Biotechnology 2 (101): 559-568 (2017)
Publisher DOI
Scopus ID
Publisher
Springer
L-tryptophan (L-trp) is a biosynthetic precursor of various bioactive components with pharmaceutical interest. The development of an efficient L-trp production strain using targeted molecular engineering approaches is challenging due to the requirement of several precursors and the complex regulations of the pathways involved. In this study, we present a rationally engineered and genetically stable L-trp overproducing Escherichia coli strain. The streamlined strain E. coli S028 is able to efficiently produce 34–40 g/L of L-trp with a yield of 0.15 g L-trp/g glucose and a productivity of 0.60 g/L/h in fed-batch fermentations. The titer and productivity of L-trp achieved are over twice as much as those reported so far for rationally developed L-trp producers. In addition, for the first time, both intracellular and extracellular concentrations of L-trp and the key metabolites in a L-trp hyperproducer strain were measured with an automated fast-sampling unit which is connected to a well-controlled bioreactor. The time series metabolic analysis gives valuable information about the regulation of L-trp synthesis in a highly productive strain and reveals targets for further improvement. Among others, it was found that L-trp and the byproduct glutamate (L-glu) accumulated to an extremely high level in the cell initially whereas the intracellular concentrations of glutamine (L-gln) stayed at a relatively low level throughout the fermentation. The metabolic analysis suggests that (a) the engineered serine biosynthesis pathway was able to effectively synthesize the substrate serine (intracellular concentration > 8 mM) for L-trp production, while (b) the substrate L-gln with an intracellular concentration of 0.8–1.2 mM seems to limit the biosynthesis of L-trp, even though L-glu was overproduced intra- and extracellularly. Thus, an increased availability of glutamine synthetase which catalyzes L-glu conversion to L-gln and an overexpression of the L-trp exporter gene could be important targets for further strain improvement.
Subjects
L-tryptophan
Rational metabolic engineering
Metabolic analysis
Escherichia coli
DDC Class
570: Biowissenschaften, Biologie
610: Medizin
620: Ingenieurwissenschaften