RoL:EAGER:DESYN-C3 Programmable Porous Lipid Sponges as Synthetic Cell Factories

RoL:EAGER:DESYN-C3 可编程多孔脂质海绵作为合成细胞工厂

• 批准号: 1844346
• 负责人: Neal Devaraj
• 金额: $ 30万
• 依托单位: University of California-San Diego
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2020-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1844346&HistoricalAwards=false
• 关键词: RoL; EAGER; DESYN; C3; Programmable

With this award, the Chemistry of Life Processes Program in the Division of Chemistry, as part of the Rules of Life (RoL): Design and Engineering of Synthetic Cells and Cell Components (DESYN-C3) initiative, is funding Dr. Neal Devaraj from the University of California, San Diego, to investigate lipid "sponges" as programmable compartments for application to the design of synthetic cells. This work capitalizes on an exciting and unexpected finding from the Devaraj lab, that biomimetic materials can be programmed to absorb biomolecules out of solution and concentrate them in a sponge-like interior. Synthetic cells have the promise to revolutionize biomanufacturing by overcoming inherent limitations faced by living cells, for instance their inability to withstand harsh conditions and toxins. Similar to organs in the human body, living cells have evolved to have interior compartments known as organelles, which have specific functions and assist in cell maintenance and function by concentrating reactants or separating mutually incompatible reactions. However it is unclear how much spatial organization and compartmentalization is necessary for the construction of a synthetic cell. Lipid sponges can help answer this question because they can be programmed to trap diverse classes of biomolecules and reactions. This project is training graduate students in supramolecular chemistry, biochemistry, soft matter, and molecular biology. The work is also contributing to outreach activities that are introducing the concept of synthetic cells to the broader San Diego educational community, with the aim of stimulating the entry of low-income and underrepresented student populations into STEM fields. The studies are providing unique insight into how compartmentalization can assist the complex chemical reactions that govern life. Living cells possess an astoundingly high macromolecular concentration. Confinement and crowding effects play critical roles in the kinetics of gene expression, protein folding, and enzymatic reactions. It has been extremely challenging to achieve reproducibly high concentrations of macromolecules inside conventional synthetic cell models such as vesicles. A lipidic mesophase system recently discovered in the Devaraj lab is being developed into a synthetic cell compartment capable of mimicking the highly crowded environment of a cell and achieving high rates and export of biological products. Surfactants can form micron-sized sponge mesophase droplets in aqueous media. These structures are termed lipid sponges to reflect their sponge-like interior network and capacity to absorb and retain biological molecules. Thanks to the high internal surface area and porous nanostructure, lipid sponges can spontaneously encapsulate high quantities of dyes and small molecules. The porous continuous structure of the lipid mesophase enables facile transport into and out of the droplets surmounting one of the key issues that has plagued lipid vesicle-based synthetic cell studies. The specific aims of this project are: (1) engineering the physico-chemical properties of lipid sponge droplets by studying the nanostructures that are formed from binary mixtures of surfactants, with the goal of improving stability and uniformity, and (2) achieving programmable compartmentalization of biochemical pathways within the droplets. Two key biochemical pathways are being studied: carbon fixation, to demonstrate the ability to supply increased levels of CO2 within a droplet and improve reaction kinetics; and controlled protein synthesis and release, through encapsulation of a DNA-programmable TX-TL (transcription-translation) system. The project has the potential to significantly advance bottom-up synthetic cell design by achieving a novel modular and programmable organelle for incorporation into artificial cells, and is providing a valuable model system for the studying the effects of colocalization and sequestration on biochemical reactions.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

With this award, the Chemistry of Life Processes Program in the Division of Chemistry, as part of the Rules of Life (RoL): Design and Engineering of Synthetic Cells and Cell Components (DESYN-C3) initiative, is funding Dr. Neal Devaraj from the University of California, San Diego, to investigate lipid "sponges" as programmable compartments for application to the design of synthetic cells.这项工作利用了Devaraj实验室的令人兴奋的意外发现，可以对仿生材料进行编程，以从溶液中吸收生物分子并将其集中在海绵状的内部中。合成细胞有望通过克服活细胞所面临的固有局限性（例如它们无法承受恶劣的条件和毒素）来彻底改变生物制造。与人体中的器官类似，活细胞已经演变为具有称为细胞器的内部隔室，它们具有特定的功能并通过浓缩反应物或分离互不兼容的反应来帮助细胞维持和功能。但是，目前尚不清楚合成细胞的构建需要多少空间组织和隔室化。脂质海绵可以帮助回答这个问题，因为它们可以编程以捕获各种类似的生物分子和反应。该项目是培训超分子化学，生物化学，软物质和分子生物学的研究生。这项工作还促进了将合成细胞概念引入更广泛的圣地亚哥教育社区的外展活动，目的是刺激低收入和代表性不足的学生人数进入STEM领域。这些研究提供了独特的见解，即分隔如何帮助控制生命的复杂化学反应。 活细胞具有惊人的高分子浓度。限制和拥挤效应在基因表达，蛋白质折叠和酶促反应的动力学中起关键作用。在常规合成细胞模型（例如囊泡）内实现可重复浓度的大分子，这是极具挑战性的。最近在Devaraj实验室中发现的脂质中间体系统正在发展成为一个合成细胞室，能够模仿细胞的高度拥挤的环境，并达到了高率和高率和生物学产品的出口。表面活性剂可以在水性培养基中形成微米大小的海绵液滴。这些结构称为脂质海绵，以反映其海绵样的内部网络和吸收和保留生物分子的能力。由于内部表面积高和多孔纳米结构，脂质海绵可以自发地封装大量的染料和小分子。脂质中间体的多孔连续结构使得易于进入和流出液滴，覆盖困扰基于脂质囊泡的合成细胞研究的关键问题之一。该项目的具体目的是：（1）通过研究由表面活性剂的二进制混合物形成的纳米结构来设计脂质海绵液滴的物理化学特性，以改善稳定性和均匀性，以及（2）实现可编程可编程的生物化学路径内部底盘的可编程范围。正在研究两种关键的生化途径：碳固定，以证明供应液滴内二氧化碳水平增加并改善反应动力学的能力；通过封装DNA可编程TX-TL（转录 - 翻译）系统，以及控制的蛋白质合成和释放。该项目有可能通过实现一种新颖的模块化和可编程细胞器来纳入人造细胞，并提供了一个有价值的模型系统，以研究共定位和隔离对生物化学反应的影响，这对NSF的法定任务和评估师的范围进行了评估。

项目成果

Lipid sponge droplets as programmable synthetic organelles

• DOI: 10.1073/pnas.2004408117
• 发表时间: 2020-08-04
• 期刊: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• 影响因子: 11.1
• 作者: Bhattacharya, Ahanjit;Niederholtmeyer, Henrike;Devaraj, Neal K.
• 通讯作者: Devaraj, Neal K.

Tailoring the Shape and Size of Artificial Cells

• DOI: 10.1021/acsnano.9b05112
• 发表时间: 2019-07-01
• 期刊: ACS NANO
• 影响因子: 17.1
• 作者: Bhattacharya, Ahanjit;Devaraj, Neal K.
• 通讯作者: Devaraj, Neal K.