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The Study of Shikimic Acid Pathway of Escherichia Coli

Author: QianJin
Tutor: ZhangHuiZhan
School: East China University of Science and Technology
Course: Biochemistry and Molecular Biology
Keywords: shikimic acid tolerance aroK gene over-expression gene-disruption
CLC: R284
Type: Master's thesis
Year: 2011
Downloads: 80
Quote: 0
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Abstract


As one of the primary intermediates in the pathway of aromatic amino acid biosynth (shikimic acid pathway), shikimic acid have series of physiological functions. And it is used as the starting material for the synthesis of the anti-H5N1 influenza virus drug under name as Tamiflu, and therefore the supply of shikimic acid is demanded.In our previous research, aroL gene which encodes shikimate kinaseⅡwas disrupte(?) Escherichia coli JM83 chromosome and the shikimic acid was accumulated in the suspen(?) of the fermentation broth. And ppsA, tktA and aroG gene were cloned from E chromosome. These genes encodes PEP synthase, transketolase and DAHP synth respectively. And the flask fermentation yield is enhanced to 94.33mg/L. In this wor fermentation conditions of pTrc-aroG-tktA/JDL02 was optimized. In 7.5h after the sec culture, using a final concentration of 0.167mg/mL of IPTG induction conditions, production of shikimic acid from pTrc-aroG-tktA/JDL02 was increase to 224.324mg/LFor investigation of the influence of the first rate-limiting enzyme of shikimic pathway, DHQ synthase which encode by aroB gene, aroB gene expressed plasmid transform into JDL02 and the flask fermentation yield was increase to 72.165mg/L. And tl aroK gene which encodes shikimate kinaseⅠwas disrupted on the chromosome Escherichia coli JM83 by means of using temperature-sensitive plasmid and addi Shikimic acid. And we confirm that inactivation of aroK would blocking up the synthesi shikimic acid by interruption of the aroB expression. In addition, we found that the shiki acid concentration on the inhibition of E. coli was between 5～10mg/mL, therefore increase of tolerance of shikimic acid would be another way to enhanced the shikimic yield.

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CLC: > Medicine, health > Chinese Medicine > Of Pharmacy > Traditional Chinese medicine chemical
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