期刊
METABOLIC ENGINEERING
卷 30, 期 -, 页码 96-104出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ymben.2015.05.001
关键词
Cytochrome P450; Synthetic biology; Plant natural products; Yeast
资金
- NIH [1S10KR02557401]
- National Institutes of Health [AT007886]
- National Science Foundation [CBET-1066100]
- Bill and Melinda Gates Foundation [OPP1058690]
- ARCS
- Div Of Chem, Bioeng, Env, & Transp Sys
- Directorate For Engineering [1066100] Funding Source: National Science Foundation
- Bill and Melinda Gates Foundation [OPP1058690] Funding Source: Bill and Melinda Gates Foundation
Microbial hosts engineered for the biosynthesis of plant natural products offer enormous potential as powerful discovery and production platforms. However, the reconstruction of these complex biosynthetic schemes faces numerous challenges clue to the number of enzymatic steps and challenging enzyme classes associated with these pathways, which can lead to issues in metabolic load, pathway specificity, and maintaining flux to desired products. Cytochrome P450 enzymes are prevalent in plant specialized metabolism and are particularly difficult to express heterologously. Here, we describe the reconstruction of the sanguinarine branch of the benzylisoquinoline alkaloid pathway in Saccharornyces cerevisine, resulting in microbial biosynthesis of protoberberine, protopine, and benzophenanthricline alkaloids through to the end-product sanguinarine, which we demonstrate can be efficiently produced in yeast in the absence of the associated biosynthetic enzyme. We achieved titers of 676 mu g/L stylopine, 548 mu g/L cis-N-methylstylopine, 252 mu g/L protopine, and 80 mu g/L sanguinarine from the engineered yeast strains. Through our optimization efforts, we describe genetic and culture strategies supporting the functional expression of multiple plant cytochrome P450 enzymes in the context of a large multi-step pathway. Our results also provided insight into relationships between cytochrome P450 activity and yeast ER physiology. We were able to improve the production of critical intermediates by 32-fold through genetic techniques and an additional 45-fold through culture optimization. (C) 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据