Engineering Yeast towards High Titer Production of Monoterpene Indole Alkaloid Natural Products

工程酵母用于高滴度生产单萜吲哚生物碱天然产物

基本信息

  • 批准号:
    10735587
  • 负责人:
  • 金额:
    $ 36.28万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
  • 财政年份:
    2018
  • 资助国家:
    美国
  • 起止时间:
    2018-07-01 至 2028-03-31
  • 项目状态:
    未结题

项目摘要

ABSTRACT Reconstruction of plant natural product pathways in genetically well-characterized microbial organisms such as Saccharomyces cerevisiae is a sustainable and scalable method of producing high value pharmaceutical compounds. The family of monoterpene indole alkaloids (MIAs) represent a diverse collection of natural product with significant biological activities. MIAs are indispensable pharmaceutical ingredients, but are also expensive and difficult to isolate from plant producers. In the previous grant cycle, we successfully engineered yeast strains that can produce strictosidine, the universal precursor to MIAs, at titers exceeding 100 mg/L. In this proposal, we will engineer the downstream steps from strictosidine to overcome key metabolic bottlenecks, and develop new yeast based-technologies for engineering heterologous natural product pathways. In collaboration with the Di Carlo lab, we will deploy PicoShell enabled cell sorting to enable high throughput screening of MIA pathways. The PicoShell technology allows microfluidic-based, high throughput single-cell encapsulation from liquid culture. Encapsulated yeast cells can be grown in bulk in a monoclonal fashion and produce the compound of interest. PicoShell effectively amplifies reporter molecule signal from single yeast cells and can be sorted with FACS based on scatter (growth rate) and fluorescence (titer). Such workflow enables the merging of yeast pathway engineering with technologies that require high throughput screening, including directed evolution and genome wide CRISPRi screening. Our preliminary efforts have shown that a fluorescent natural product in the MIA pathway can serve as a reporter for the efficiency of the downstream steps during PicoShell enabled FACS sorting. This collaborative proposal will leverage Tang lab’s expertise in natural product biosynthesis with the new nanobiotechnology tools developed for yeast by the Di Carlo Lab. This will pave the way for complete reconstitution of important MIAs at high titers in yeast, as well as establishing new tools for yeast synthetic biology. Together we will address three aims: 1) overcoming key bottleneck step in post- strictosidine steps, specifically the low efficiency of strictosidine glucosidase (SGD); 2) host engineering with CRISPR interference and activation to increase strictosidine levels, using both rational and genome wide screening enabled by PicoShells; and 3) complete biosynthesis of complex MIAs ibogaine and mitragynine, two psychoactive MIAs that have generated significant interests as potential treatment for opioid addiction.
摘要

项目成果

期刊论文数量(0)
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Dino Di Carlo其他文献

Dino Di Carlo的其他文献

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{{ truncateString('Dino Di Carlo', 18)}}的其他基金

Hydrogel nanovial technology for single-cell sorting based on extracellular vesicle production
基于细胞外囊泡产生的单细胞分选水凝胶纳米瓶技术
  • 批准号:
    10411907
  • 财政年份:
    2021
  • 资助金额:
    $ 36.28万
  • 项目类别:
Enhancing the potency of mesenchymal stem cell therapies for kidney diseases using lab-on-a-particle technology
使用粒子实验室技术增强间充质干细胞治疗肾脏疾病的效力
  • 批准号:
    10373803
  • 财政年份:
    2021
  • 资助金额:
    $ 36.28万
  • 项目类别:
Lab on a particle technology for functional screening of therapeutic cells
用于治疗细胞功能筛选的粒子技术实验室
  • 批准号:
    10272940
  • 财政年份:
    2021
  • 资助金额:
    $ 36.28万
  • 项目类别:
Hydrogel nanovial technology for single-cell sorting based on extracellular vesicle production
基于细胞外囊泡产生的单细胞分选水凝胶纳米瓶技术
  • 批准号:
    10193200
  • 财政年份:
    2021
  • 资助金额:
    $ 36.28万
  • 项目类别:
Caltech/UCLA Individualized Theranostic Engineering to Advance Metabolic System (iTEAM)
加州理工学院/加州大学洛杉矶分校个性化治疗诊断工程促进代谢系统 (iTEAM)
  • 批准号:
    10213026
  • 财政年份:
    2020
  • 资助金额:
    $ 36.28万
  • 项目类别:
Caltech/UCLA Individualized Theranostic Engineering to Advance Metabolic System (iTEAM)
加州理工学院/加州大学洛杉矶分校个性化治疗诊断工程促进代谢系统 (iTEAM)
  • 批准号:
    10440285
  • 财政年份:
    2020
  • 资助金额:
    $ 36.28万
  • 项目类别:
Caltech/UCLA Individualized Theranostic Engineering to Advance Metabolic System (iTEAM)
加州理工学院/加州大学洛杉矶分校个性化治疗诊断工程促进代谢系统 (iTEAM)
  • 批准号:
    10683974
  • 财政年份:
    2020
  • 资助金额:
    $ 36.28万
  • 项目类别:
Training the next generation of leaders in biomedical engineering design
培训下一代生物医学工程设计领导者
  • 批准号:
    10599275
  • 财政年份:
    2019
  • 资助金额:
    $ 36.28万
  • 项目类别:
Training the next generation of leaders in biomedical engineering design
培训下一代生物医学工程设计领导者
  • 批准号:
    10428473
  • 财政年份:
    2019
  • 资助金额:
    $ 36.28万
  • 项目类别:
Force phenotyping of airway smooth muscle cells to develop novel asthma therapies
强制气道平滑肌细胞表型分析以开发新型哮喘疗法
  • 批准号:
    9452964
  • 财政年份:
    2017
  • 资助金额:
    $ 36.28万
  • 项目类别:

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骨骼合成代谢过程中骨-脂肪相互作用
  • 批准号:
    10590611
  • 财政年份:
    2022
  • 资助金额:
    $ 36.28万
  • 项目类别:
Bone-Adipose Interactions During Skeletal Anabolism
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Bone-Adipose Interactions During Skeletal Anabolism
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  • 批准号:
    10365254
  • 财政年份:
    2021
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    $ 36.28万
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Bone-Adipose Interactions During Skeletal Anabolism
骨骼合成代谢过程中骨-脂肪相互作用
  • 批准号:
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BCCMA: Foundational Research to Act Upon and Resist Conditions Unfavorable to Bone (FRACTURE CURB): Combined long-acting PTH and calcimimetics actions on skeletal anabolism
BCCMA:针对和抵抗不利于骨骼的条件的基础研究(遏制骨折):长效 PTH 和拟钙剂联合作用对骨骼合成代谢的作用
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Promotion of NAD+ anabolism to promote lifespan
促进NAD合成代谢以延长寿命
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    DE170100628
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    2017
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