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Optimization of Baccatin III Production by Cross-Linked Enzyme Aggregate of Taxoid 10β-O-Acetyltransferase

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Abstract

Taxoid 10β-O-acetyltransferase (DBAT) is the key enzyme to produce baccatin III, a key precursor in paclitaxel synthesis, by acetyl group transfer from acetyl-CoA to the C10 hydroxyl of 10-deacetylbaccatin III. In this study, the recombinant DBAT (rDBAT) was immobilized by cross-linked enzyme aggregates (CLEAs). To further optimize the enzyme recovery, single-factor experiment and response surface methodology were applied. 60% ammonium sulfate as precipitant, 0.05% glutaraldehyde as fixing agent, pH 7.0, 2 h as cross-linking time, 30 °C as cross-linking temperature were confirmed to be the optimum conditions to prepare the CLEAs-rDBAT in single-factor experiment. In addition, 62% for ammonium sulfate saturation, 0.15% for glutaraldehyde, and pH 6.75 were confirmed to be the optimum conditions with averagely 73.9% activity recovery in 3 replications, which was consistent with the prediction of response surface methodology. After cross-linking, the optimum temperature of CLEAs-rDBAT rose up to 70 °C and CLEAs-rDBAT could be recycled for three times.

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Acknowledgements

This work was supported by the Science and Technology Program of Guangdong Province (Grants 2014B050505018, 2014B020205003, 2015A020209121) and the National Natural Science Foundation of China (Grants 31071837, 31572178, 31772373).

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Author notes

  1. Lin-Feng You and Tao Wei are co-first authors. They contributed equally to this work.

Authors and Affiliations

  1. College of Food Science, Institute of Food Biotechnology, South China Agricultural University, Guangzhou, 510640, Guangdong, China

    Lin-Feng You, Tao Wei, Qian-Wang Zheng, Jun-Fang Lin & Li-Qiong Guo

  2. Chongqing Key Laboratory of Catalysis and Functional Organic Molecule, Chongqing Technology and Business University, Chongqing, 400067, China

    Lin-Feng You

  3. Research Center for Micro-Ecological Agent Engineering and Technology of Guangdong Province, Guangzhou, 510640, Guangdong, China

    Tao Wei, Qian-Wang Zheng, Jun-Fang Lin & Li-Qiong Guo

  4. Department of Pathology, Anatomy and Cell Biology, Thomas Jefferson University, Philadelphia, PA, 19107, USA

    Bing-Hua Jiang

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  1. Lin-Feng You
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Correspondence to Jun-Fang Lin or Li-Qiong Guo.

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You, LF., Wei, T., Zheng, QW. et al. Optimization of Baccatin III Production by Cross-Linked Enzyme Aggregate of Taxoid 10β-O-Acetyltransferase. Mol Biotechnol 61, 498–505 (2019). https://doi.org/10.1007/s12033-019-00179-1

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  • DOI: https://doi.org/10.1007/s12033-019-00179-1

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