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Hydroaromatic equilibration during biosynthesis of shikimic acid
D R Knop1, K M Draths, S S Chandran
1The Department of Chemistry, Michigan State University, East Lansing, Michigan 48824-1322, USA.
Journal of the American Chemical Society
|October 18, 2001
Summary
Improving microbial production of shikimic acid, a valuable synthetic precursor, requires minimizing byproduct formation. Researchers identified that quinic acid byproduct formation stems from shikimic acid transport and equilibration within Escherichia coli.
Area of Science:
- Biotechnology
- Metabolic Engineering
- Microbial Synthesis
Background:
- Shikimic acid is a crucial hydroaromatic compound for chemical synthesis, but its plant-derived isolation is expensive and limited.
- Recombinant Escherichia coli strains have been developed for shikimic acid biosynthesis from glucose, yet yields are often reduced by significant byproduct formation, primarily quinic acid and 3-dehydroshikimic acid.
- Understanding the mechanisms behind byproduct formation is essential for optimizing shikimic acid production.
Purpose of the Study:
- To elucidate the mechanism of quinic acid formation during microbial shikimic acid biosynthesis.
- To identify strategies for reducing quinic acid and improving the overall yield and purity of synthesized shikimic acid.
- To enhance the efficiency of Escherichia coli as a cell factory for shikimic acid production.
Main Methods:
- Utilized recombinant Escherichia coli strains (E. coli SP1.1/pKD12.138 and E. coli SP1.1/pSC5.214A) engineered for shikimic acid synthesis.
- Investigated byproduct formation by analyzing the molar ratios of shikimate, quinate, and dehydroshikimate under different conditions.
- Performed experiments with a strain incapable of de novo shikimic acid synthesis to study the conversion of exogenous shikimic acid into byproducts.
- Manipulated shikimate transport to assess its impact on quinic acid formation.
Main Results:
- The typical synthesis of shikimic acid by E. coli yielded a mixture containing significant amounts of quinic acid and 3-dehydroshikimic acid.
- Experiments demonstrated that quinic acid formation is linked to the microbe-catalyzed equilibration of initially synthesized shikimic acid, likely involving cytoplasmic transport.
- Repressing shikimate transport in E. coli SP1.1/pKD12.138 significantly increased shikimic acid titer to 52 g/L with improved purity (14:1.0:3.0 shikimate/quinate/dehydroshikimate ratio).
Conclusions:
- Quinic acid byproduct formation during microbial shikimic acid biosynthesis is primarily due to an equilibration process involving shikimic acid transport and subsequent metabolic reactions within the cell.
- Engineering strategies that control or repress shikimate transport can substantially enhance the yield and purity of biosynthesized shikimic acid.
- This study provides critical mechanistic insights for optimizing metabolic engineering strategies to improve the industrial production of shikimic acid using Escherichia coli.
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