Walls, LE;
Malcı, K;
Nowrouzi, B;
Li, RA;
d'Espaux, L;
Wong, J;
Dennis, JA;
... Rios-Solis, L; + view all
(2021)
Optimizing the biosynthesis of oxygenated and acetylated Taxol precursors in Saccharomyces cerevisiae using advanced bioprocessing strategies.
Biotechnology and Bioengineering
, 118
(1)
pp. 279-293.
10.1002/bit.27569.
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Abstract
Taxadien-5α-hydroxylase and taxadien-5α-ol O-acetyltransferase catalyze the oxidation of taxadiene to taxadien-5α-ol and subsequent acetylation to taxadien-5α-yl-acetate in the biosynthesis of the blockbuster anticancer drug, paclitaxel (Taxol®). Despite decades of research, the promiscuous and multispecific CYP725A4 enzyme remains a major bottleneck in microbial biosynthetic pathway development. In this study, an interdisciplinary approach was applied for the construction and optimization of the early pathway in Saccharomyces cerevisiae, across a range of bioreactor scales. High-throughput microscale optimization enhanced total oxygenated taxane titer to 39.0 ± 5.7 mg/L and total taxane product titers were comparable at micro and minibioreactor scale at 95.4 ± 18.0 and 98.9 mg/L, respectively. The introduction of pH control successfully mitigated a reduction of oxygenated taxane production, enhancing the potential taxadien-5α-ol isomer titer to 19.2 mg/L, comparable with the 23.8 ± 3.7 mg/L achieved at microscale. A combination of bioprocess optimization and increased gas chromatography-mass spectrometry resolution at 1 L bioreactor scale facilitated taxadien-5α-yl-acetate detection with a final titer of 3.7 mg/L. Total oxygenated taxane titers were improved 2.7-fold at this scale to 78 mg/L, the highest reported titer in yeast. Critical parameters affecting the productivity of the engineered strain were identified across a range of scales, providing a foundation for the development of robust integrated bioprocess control systems.
| Type: | Article |
|---|---|
| Title: | Optimizing the biosynthesis of oxygenated and acetylated Taxol precursors in Saccharomyces cerevisiae using advanced bioprocessing strategies |
| Location: | United States |
| Open access status: | An open access version is available from UCL Discovery |
| DOI: | 10.1002/bit.27569 |
| Publisher version: | https://doi.org/10.1002/bit.27569 |
| Language: | English |
| Additional information: | Copyright © 2020 The Authors. Biotechnology and Bioengineering published by Wiley Periodicals LLC. This is an open access article under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits use, distribution and reproduction in any medium, provided the original work is properly cited. |
| Keywords: | Saccharomyces cerevisiae, Taxol, high throughput microbioreactor, taxadien-5-alpha-olO-acetyltransferase, taxadien-5-hydroxylase, Bridged-Ring Compounds, Metabolic Engineering, Paclitaxel, Saccharomyces cerevisiae, Taxoids |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > UCL BEAMS UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Engineering Science UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Engineering Science > Dept of Biochemical Engineering |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10183108 |
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