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A Customizable Approach for the Enzymatic Production and Purification of Diterpenoid Natural Products
Published on: October 4, 2019
Terrequinone A biosynthesis through L-tryptophan oxidation, dimerization and bisprenylation
Carl J Balibar1, Annaleise R Howard-Jones, Christopher T Walsh
1Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Ave., Boston, Massachusetts 02115, USA.
Researchers elucidated the biosynthesis of the antitumor compound terrequinone A. They reconstituted the pathway in E. coli, revealing key enzymatic steps including L-tryptophan aminotransferase and prenyltransferase activities.
Area of Science:
- Biochemistry
- Natural Product Biosynthesis
- Molecular Biology
Background:
- Terrequinone A is an antitumor fungal metabolite produced by Aspergillus sp.
- Its biosynthesis is governed by the five-gene cluster tdiA-tdiE.
Purpose of the Study:
- To overproduce and biochemically characterize the proteins involved in terrequinone A biosynthesis.
- To elucidate the complete pathway, including enzymatic mechanisms and intermediates.
Main Methods:
- Overproduction of terrequinone A biosynthetic proteins (TdiA-TdiE) in Escherichia coli.
- Biochemical assays to determine the function of individual enzymes.
- Characterization of reaction intermediates and products.
Main Results:
- Reconstitution of terrequinone A biosynthesis in E. coli.
- Identification of TdiD as an L-tryptophan aminotransferase generating indolepyruvate.
- Characterization of TdiA as a tridomain nonribosomal peptide synthetase involved in nonoxidative dimerization.
- TdiC identified as an NADH-dependent quinone reductase facilitating prenylation.
- TdiB confirmed as the prenyltransferase responsible for bisprenylation.
- TdiE's role in preventing off-pathway products elucidated.
Conclusions:
- The complete biosynthetic pathway of terrequinone A has been biochemically characterized.
- The pathway involves novel nonoxidative dimerization and asymmetric prenylation steps.
- Understanding this pathway provides insights into natural product biosynthesis and potential for synthetic biology applications.
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