Refactoring Soft Coral Diterpenoid Biosynthesis
重构软珊瑚二萜生物合成
基本信息
- 批准号:10553605
- 负责人:
- 金额:$ 7.18万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2022
- 资助国家:美国
- 起止时间:2022-02-01 至 2025-01-31
- 项目状态:未结题
- 来源:
- 关键词:AnabolismAquacultureArchitectureBindingBiochemistryBioinformaticsBiologicalBiological AssayCarbonCell LineCharacteristicsChemicalsChemistryCloningCommon CoreComplexCyclobutanesCytochrome P450CytochromesDataDevelopmentDirected Molecular EvolutionDiterpenesEngineeringEnzymatic BiochemistryEnzymesEukaryotaEvaluationEventExhibitsFamilyFutureGene ClusterGenesGenetic TranscriptionGenomeGeranylgeranyl-diphosphate geranylgeranyltransferaseGoalsHandHumanHydrocarbonsIn VitroK-562LeadLeukemic CellLyaseMedicineMembraneMethodsMutagenesisNatural ProductsNatural SourceOxidasesOxidoreductasePathway interactionsPeripheralPharmacologic SubstancePichiaProductionPublicationsPublishingReactionReportingResearchSeriesSourceStructureTechniquesTerpenesTerpenoid Biosynthesis PathwayTherapeuticVariantVertebral columnWorkYeastsanti-cancercombinatorialcoralcytotoxicitydesign and constructiondesign,build,testenzyme biosynthesisimprovedin vivoinsightlycopenemarinemarine natural productmicrobialmodel developmentnoveloxidationpromoterrapid detectionreconstitutionresearch clinical testingscaffoldsecondary metaboliteterpene synthasetranscriptome
项目摘要
Project Summary
Soft corals (Alcyonaria) are a remarkable source of natural products (NPs) with pharmaceutically relevant
biological activity, and complex scaffolds that are unique to marine eukaryotes with no terrestrial sources of
structurally related compounds. The composition of secondary metabolites isolated from soft corals is dominated
by a diverse suite of bioactive diterpenes with over 50 unique scaffolds and >1500 unique diterpenoids isolated
to date. Many coral-derived diterpenes have potent and selective biological activity, including the xenicane,
acalycixeniolide F, which displays cytotoxicity against human leukemia cell line K562 (LC50 = 200 ng/mL) via an
undetermined mode of action. However, despite the promise these compounds exhibit as lead structures for the
development of novel medicines, the pharmaceutical potential of coral diterpenes remains untapped as neither
total synthesis nor isolation from aquaculture have provided sufficient material to enable effective in vivo clinical
testing. Through bioinformatic analysis of published soft coral genomes and transcriptomes, our group has
recently identified a series of terpene synthases that synthesize the hydrocarbon backbones of multiple coral-
specific diterpenes, including xeniaphyllene, the precursor to the archetypal coral diterpenes, the xenicanes.
Here, I propose to use the biosynthesis of the xenicanes as model for the development of a sustainable platform
for the microbial production of bioactive coral diterpenes.
This proposal aims to optimize the production of xenicane diterpenes through characterization and
directed evolution of enzymes involved in the synthesis and oxidation of the xenicane scaffold, culminating in
the reconstitution of xenicane biosynthesis through pathway engineering in methylotrophic yeast, Pichia pastoris.
Terpene synthases are often the rate-limiting step in terpenoid biosynthesis due to their low catalytic efficiency
and instability. In Aim 1, I will use a high-throughput, colorimetric substrate competition assay to screen error-
prone PCR-generated xeniaphyllene terpene synthase variants for improved activity and stability. To access the
core scaffold common to all xenicanes, the fused cyclobutane ring in xeniaphyllene must undergo an oxidative
carbon-carbon bond cleavage. In the recently published chromosomal level genome assembly of a Xenia sp.,
we found a suite of cytochromes P450 co-localized with the xeniaphyllene synthase. In Aim 2, I will explore the
oxidative chemistry of these cytochromes P450 in search of the unprecedented carbon–carbon lyase. Finally,
the reconstitution of xenicane biosynthesis will be explored in Aim 3, wherein I will use Universal Loop Assembly
(uLoop) to efficiently design and construct a refactored biosynthetic pathway for fermentative production of
xenicanes in P. pastoris. This research will provide invaluable insights into the enzymology behind the
construction of the biologically active xenicane coral diterpenes. Furthermore, the methods developed from this
work will facilitate the reconstitution of additional marine metazoan biosynthetic pathways, resolving supply
issues for further biological evaluation of these NPs.
项目摘要
软珊瑚(Alcyonaria)是天然产物(NPs)的重要来源,具有药用价值
生物活性,以及没有陆地来源的海洋真核生物所独有的复杂支架
结构上相关的化合物。从软珊瑚中分离出的次生代谢物的组成占主导地位。
由一套不同的生物活性二萜类化合物组成,具有50多种独特的支架和1500种独特的二萜类化合物
到目前为止。许多珊瑚衍生的二萜化合物具有强大的和选择性的生物活性,包括十一烷,
对人白血病细胞株K562(LC_(50)=200 ng/mL)的细胞毒作用。
未确定的行动模式。然而,尽管这些化合物显示出作为先导结构的前景
新药的开发,珊瑚二萜类化合物的药用潜力仍未开发,因为
从水产养殖中完全合成或分离都提供了足够的材料来实现有效的活体临床
测试。通过对已发表的软珊瑚基因组和转录本进行生物信息学分析,我们小组已经
最近发现了一系列合成多个珊瑚碳氢化合物骨架的萜烯合成酶-
特定的二萜类化合物,包括原型珊瑚二萜类化合物的前体--香叶烯。
在这里,我建议将异构烷的生物合成作为可持续平台开发的模型
用于微生物生产生物活性珊瑚二萜。
这项建议旨在通过表征和分析来优化十二烷二萜的生产
参与合成和氧化硅烷支架的酶的定向进化,最终达到
甲醇酵母巴斯德毕赤酵母通过途径工程重组异环己烷生物合成。
萜烯合酶由于催化效率低,常常是萜类化合物生物合成的限速步骤。
和不稳定。在目标1中,我将使用高通量、比色底物竞争分析来筛选错误-
容易聚合酶链式反应产生的异枝烯萜烯合酶变种,以提高活性和稳定性。要访问
所有杂环烷共有的核心支架,异构烯中稠合的环丁烷环必须经历氧化
碳-碳键断裂。在最近发表的一株Xenia sp.的染色体水平基因组组装中,
我们发现了一组细胞色素P450,它们与异烯合成酶共定位。在目标2中,我将探索
这些细胞色素P450的氧化化学寻找前所未有的碳-碳裂解酶。最后,
在目标3中,我将使用万能环路组件来探索异烷生物合成的重组
(ULoop)高效设计和构建用于发酵生产的可重构生物合成途径
巴氏拟青霉中的二十二烷类化合物。这项研究将提供宝贵的见解,以了解背后的酶
具有生物活性的珊瑚二萜化合物的构建。此外,在此基础上发展起来的方法
工作将促进额外的海洋后生动物生物合成途径的重建,解决供应
关于进一步对这些NPs进行生物学评估的问题。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
数据更新时间:{{ journalArticles.updateTime }}
{{
item.title }}
{{ item.translation_title }}
- DOI:
{{ item.doi }} - 发表时间:
{{ item.publish_year }} - 期刊:
- 影响因子:{{ item.factor }}
- 作者:
{{ item.authors }} - 通讯作者:
{{ item.author }}
数据更新时间:{{ journalArticles.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ monograph.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ sciAawards.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ conferencePapers.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ patent.updateTime }}
Vikram Vijay Shende其他文献
Vikram Vijay Shende的其他文献
{{
item.title }}
{{ item.translation_title }}
- DOI:
{{ item.doi }} - 发表时间:
{{ item.publish_year }} - 期刊:
- 影响因子:{{ item.factor }}
- 作者:
{{ item.authors }} - 通讯作者:
{{ item.author }}
{{ truncateString('Vikram Vijay Shende', 18)}}的其他基金
相似海外基金
Development of a mucosally-delivered and active salmon louse vaccine for Atlantic salmon aquaculture
开发用于大西洋鲑鱼水产养殖的粘膜递送活性鲑鱼虱疫苗
- 批准号:
BB/Y006534/1 - 财政年份:2024
- 资助金额:
$ 7.18万 - 项目类别:
Research Grant
Protecting oyster aquaculture from heatwaves and flooding rains
保护牡蛎养殖免受热浪和洪水的影响
- 批准号:
DE240100272 - 财政年份:2024
- 资助金额:
$ 7.18万 - 项目类别:
Discovery Early Career Researcher Award
Intelligent Dependable Environment Control For Sustainable Aquaculture
可持续水产养殖的智能可靠环境控制
- 批准号:
EP/Y000773/1 - 财政年份:2024
- 资助金额:
$ 7.18万 - 项目类别:
Research Grant
AQUAWAVE: Advancing Quality and Understanding of Wave Conditions in new Aquaculture Environments
AQUAWAVE:提高新水产养殖环境中波浪条件的质量和理解
- 批准号:
NE/Y005198/1 - 财政年份:2024
- 资助金额:
$ 7.18万 - 项目类别:
Research Grant
Advancing Shrimp Farming: Environmental DNA and RNA as biomarkers in shrimp aquaculture
推进虾类养殖:环境 DNA 和 RNA 作为虾类水产养殖的生物标志物
- 批准号:
24K09059 - 财政年份:2024
- 资助金额:
$ 7.18万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
I-Corps: High Intensity pulsed light for the removal of off-flavor compounds in aquaculture effluents
I-Corps:高强度脉冲光用于去除水产养殖废水中的异味化合物
- 批准号:
2306173 - 财政年份:2023
- 资助金额:
$ 7.18万 - 项目类别:
Standard Grant
Integrating food and nutrition into fisheries and aquaculture management
将粮食和营养纳入渔业和水产养殖管理
- 批准号:
DE230100069 - 财政年份:2023
- 资助金额:
$ 7.18万 - 项目类别:
Discovery Early Career Researcher Award
Collaborative Research: Management Practices and Microbial Communities in Prehistoric Aquaculture
合作研究:史前水产养殖的管理实践和微生物群落
- 批准号:
2242083 - 财政年份:2023
- 资助金额:
$ 7.18万 - 项目类别:
Standard Grant
Bringing the phenomics of aquatic embryos to key challenges in aquaculture
将水生胚胎的表型组学应用于水产养殖的关键挑战
- 批准号:
BB/W017938/1 - 财政年份:2023
- 资助金额:
$ 7.18万 - 项目类别:
Research Grant
A Behavioral Economics Approach to Sustainable Aquaculture Business: Social Experiments in Developing Countries
可持续水产养殖业务的行为经济学方法:发展中国家的社会实验
- 批准号:
23KK0029 - 财政年份:2023
- 资助金额:
$ 7.18万 - 项目类别:
Fund for the Promotion of Joint International Research (International Collaborative Research)