Abstract
L-tyrosine is a commercially important compound in the food, pharmaceutical, chemical, and cosmetic industries. Although several attempts have been made to improve L-tyrosine production, translation-level expression control and carbon flux rebalancing around phosphoenolpyruvate (PEP) node still remain to be achieved for optimizing the pathway. Here, we demonstrate pathway optimization by altering gene expression levels for L-tyrosine production in Escherichia coli. To optimize the L-tyrosine biosynthetic pathway, a synthetic constitutive promoter and a synthetic 5′-untranslated region (5′-UTR) were introduced for each gene of interest to allow for control at both transcription and translation levels. Carbon flux rebalancing was achieved by controlling the expression level of PEP synthetase using UTR Designer. The L-tyrosine productivity of the engineered E. coli strain was increased through pathway optimization resulting in 3.0 g/L of L-tyrosine titer, 0.0354 g L-tyrosine/h/g DCW of productivity, and 0.102 g L-tyrosine/g glucose yield. Thus, this work demonstrates that pathway optimization by 5′-UTR redesign is an effective strategy for the development of efficient L-tyrosine-producing bacteria.
Highlights
L-tyrosine is a commercially important compound in the food, pharmaceutical, chemical, and cosmetic industries
If carbon flux at the phosphoenolpyruvate (PEP) node was increased toward the L-tyrosine biosynthetic pathway through genetic engineering, carbon flux to the tricarboxylic acid (TCA) cycle would be decreased
In E. coli, the elements of the L-tyrosine synthetic pathway are encoded by eleven genes
Summary
In E. coli, the elements of the L-tyrosine synthetic pathway are encoded by eleven genes. In the tyrosine synthetic pathway, AroG, AroB, and TyrA are negatively regulated by L-tyrosine[41,42]. The TyrR protein, a regulator expressed in response to intracellular tyrosine concentrations, tightly regulates the expression of genes such as AroL, TyrA, and TyrB13,43. A 5′-UTR sequence was designed by UTR Designer to achieve the maximum expression level of each gene in the tyrosine pathway (Fig. 1a). Based on the N-terminal coding sequence of each gene, UTR Designer generated different 5′ -UTR sequences that can obtain maximum expression levels (Supplementary Table S1). Since the 5′ -UTR sequences can be different depending on coding sequences that we input, we selected the sequence with the highest expression level for each gene and replaced it
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