Abstract
ABSTRACT Tryptophan, an aromatic amino acid, has been widely used in food industry because it participates in the regulation of protein synthesis and metabolic network in vivo. In this study, we obtained a strain named TRP03 by enhancing the tryptophan synthesis pathway, which could accumulate tryptophan at approximately 35 g/L in a 5 L bioreactor. We then modified the central metabolic pathway of TRP03, to increase the supply of the precursor phosphoenolpyruvate (PEP), the genes related to PEP were modified. Furthermore, citric acid transport system and TCA were upregulated to effectively increase cell growth. We observed that strain TRP07 that could accumulate tryptophan at approximately 49 g/L with a yield of 0.186 g tryptophan/g glucose in a 5 L bioreactor. By-products such as glutamate and acetic acid were reduced to 0.8 g/L and 2.2 g/L, respectively.
Highlights
Tryptophan, an aromatic amino acid [1], plays an important role in various biological activities in living beings including humans
Microorganisms commonly used in tryptophan production include Escherichia coli and Corynebacterium glutamiens, Corynebacterium glutamatum is a natural microorganism for producing amino acids [11], E. coli (Escherichia coli), which has a faster reproductive cycle and more mature genetic modification systems, has been widely applied to the microbial fermentation industry, including the production of amino acids, such as tryptophan, threonine [12], valine [13], tyrosine [14]; organic acids, such as malic acid [15], succinic acid [16], lactic acid [17], glutaric acid [18], and adipic acid [19], among others
Downregulation of the tryptophan degradation pathway and transport system Mtr, TnaB, and AroP are the three transporters in Escherichia coli related to L-tryptophan
Summary
Tryptophan, an aromatic amino acid [1], plays an important role in various biological activities in living beings including humans. L-tryptophan is produced by extraction, chemical synthesis [7,8], enzymatic, and microbial fermentation methods [9]. In E. coli, tryptophan is metabolized [22] with the help of the glycolytic, pentose phosphate, shikimic acid, and chorismate pathways. PEP and erythrose-4-phosphate (E4P), as carbon skeleton precursors directly enter the shikimic acid pathway, are the most important precursors for tryptophan synthesis. The main function of PEP is to enter the TCA cycle via pyruvate to provide energy for cell growth, and the proportion of carbon metabolic flow into the shikimic acid pathway usually does not exceed 2% [31]. Weakening the PEP-to-pyruvate pathway to enhance carbon flux of the tryptophan biosynthesis pathway affects cell growth [34]. The titer of tryptophan was further improved to 49 g/L, which is 40% higher than 35 g/L, with a yield of 0.186 g tryptophan/g glucose in a 5 L bioreactor
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