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| Effects of Modification in Operon-leader Region and Strengthening in PRPP Synthesis Pathway on L-histidine Accumulation by Escherichia coli |
| LU Li-ning, CHENG Yong-song, XIE Xi-xian, XU Qing-yang, CHEN Ning |
| College of Biological Engineering, Tianjin University of Science & Technology, Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin 300457, China |
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Abstract Objective: Based on Escherichia coli MG1655-ΔtktA, histidine-operon leader region was replaced by strong promoter T5, 6-phosphoglucose dehydrogenase(zwf), 6-phosphogluconate dehydrogenase(gnd) and phosphoribosylpyrophosphate synthetase(prs) were overexpressed. Effects of the above modifications on L-histidine accumulation were investigated. Methods: In strain with tktA interrupted, leader region in histidine operon was replaced by T5 promoter by Red recombination system from bacteriophage λ. By cloning technology, zwf and gnd were coexpressed on pSTV28 and prs was expressed on pQE30. Effects of the above modifications on L-histidine accumulation were compared by fermentation in flasks. Results: Quantified by HPLC, trace L-histidine was accumulated in fermentation mediums for those strains with leader region replaced by promoter T5——MG1655-ΔtktA-PT5, MG1655-ΔtktA-PT5(prs-pQE30), MG1655-ΔtktA-PT5(zwf-gnd-pSTV28), and MG1655-ΔtktA-PT5 (prs-pQE30, zwf-gnd-pSTV28) with 60.12 mg/L, 66.47 mg/L, 89.69 mg/L and 111.56 mg/L, respectively. Conclusion: Modification of leader region in operon strengthened the ability to synthesize L-histidine. Afterward, elevation in oxidative pentose phosphate pathway level and PRPP synthetase activity resulted in increased accumulation.
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Received: 05 December 2011
Published: 25 April 2012
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