Engineered Consortia for Effective Utilization of a Cyanobacterial Carbohydrate Feedstock

有效利用蓝藻碳水化合物原料的工程联合体

• 批准号: 1437657
• 负责人: Daniel Ducat
• 金额: $ 29.98万
• 依托单位: Michigan State University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2018-02-28
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1437657&HistoricalAwards=false
• 关键词: Engineered; Consortia; Effective; Utilization; Cyanobacterial

Principal Investigator: Daniel DucatNumber: 1437657Currently, the production of biofuels relies on carbohydrate feedstocks sourced from plant-based crop species that compete with edible crop species for arable landmass and potable water. Alternative, photosynthetically derived feedstock sources of carbohydrates can uncouple biofuel production from traditional agriculture and potentially decrease the land footprint required. This project will make use of a genetically engineered strain of a photosynthetic bacterium (cyanobacterium) that will use carbon dioxide in the atmosphere and light to make sucrose (table sugar), which can be fermented to biofuels and bio-products. This microorganism is uniquely capable of rerouting the majority of its photosynthetically fixed carbon towards the production and excretion of sucrose. This engineered cyanobacterium has carbohydrate productivity potential that exceeds that of traditional plant species, and could be used for an alternative carbohydrate feedstock if effective approaches can be designed to scale culture growth and minimize the cost of feedstock recovery. The project will focus on the development of microbial consortia as one means to capitalize upon the productive capacity of these sucrose-secreting cyanobacteria. In this approach, the sugar-secreting cyanobacterium with will be co-cultured with yeast strains that efficiently utilize the carbohydrates and biologically convert them into commodity products without the requirement of sucrose purification or processing. Improvements in the capacity for the yeast stain to robustly grow and utilize cyanobacterial sugars will also ultimately improve the conversion of sucrose into biofuels and bioproducts. Towards this end, a model microbial consortium will be developed whereby engineered yeast will utilize photosynthetically produced sucrose for the production of fatty alcohols, a useful surfactant and potential biofuel additive. This project includes research and educational experiences for a graduate student, as well as undergraduate students through the Plant Genomics Program and Khorana Scholars Program at Michigan State University. In addition, this project involves engagement with an undergraduate research program at Schoolcraft College, and science policy discussions on the use of synthetic biology for biotechnology applications through a Sloan Institute sponsored Delphi policy study. Technical DescriptionThe overall goal of this project is to develop a co-culture system for the overproduction of sucrose from carbon dioxide and light using engineered strains of cyanobacteria and yeast. The approach relies on a two-step photobiological production of sucrose through co-culture, and will provide fundamental understanding on how engineer of inter-species interactions for this purpose. This project will make use of an engineered cyanobacterial strain, Synechococcus elongatus PCC 7942 (S. elongatus) recently developed in the PI?s laboratory that is genetically stable and capable of rerouting up to 85% of its fixed carbon towards excreted sucrose. This strain possesses a high specific activity for photosynthetic production and therefore has considerable potential as an alternative carbohydrate feedstock source if scaled culturing can mitigate three challenges, which will be addressed by the proposed research. These challenges are to 1) minimize reactor complexity and cost to a degree consistent with the commodity product value, 2) reduce the costs of purifying, processing, and delivering cyanobacterial carbohydrates to downstream applications, and 3) to reduce the risk of photobioreactor contamination. Towards this end, the project will focus on the development of a direct co-culture of S. elongates with Saccharomyces cerevisiae (yeast) that will flexibly convert the sucrose into added-value compounds. The project will explore specific strategies to improve yeast growth in co-culture, including, improvement of yeast carbon uptake, increased yeast tolerance of cyanobacterial secondary products of photosynthetic metabolism, and enhancement of the flux and specificity of carbon from cyanobacteria to yeast through engineered physical association. These strategies will be evaluated to improve yeast robustness and efficiency of conversion of carbohydrates derived from S. elongatus when grown in co-culture, and a final proof-of-principle production of cetyl alcohol is proposed through the use of an established fatty-alcohol producing yeast strain. Overall, the project has potential to impact the field of algal biofuels by providing an alternative approach for metabolic pathway engineering or by improving upon an attractive algae-derived feedstock. This project includes research and educational experiences for a graduate student, as well as undergraduates through the Plant Genomics Program and Khorana Scholars Program at Michigan State University. In addition, this project involves engagement with an undergraduate research program at Schoolcraft College, and science policy discussions on the use of synthetic biology for biotechnology applications through a Sloan Institute sponsored Delphi policy study.

首席研究员：Daniel Ducat 编号：1437657 目前，生物燃料的生产依赖于源自植物作物的碳水化合物原料，这些作物与可食用作物争夺耕地和饮用水。替代性的、光合作用衍生的碳水化合物原料来源可以将生物燃料生产与传统农业脱钩，并有可能减少所需的土地足迹。该项目将利用光合细菌（蓝藻）的基因工程菌株，利用大气中的二氧化碳和光来制造蔗糖（食糖），蔗糖可以发酵成生物燃料和生物产品。 这种微生物具有独特的能力，能够将其大部分光合固定碳重新用于蔗糖的生产和排泄。这种工程蓝细菌具有超过传统植物物种的碳水化合物生产力潜力，如果可以设计有效的方法来扩大培养物生长并最大限度地降低原料回收成本，则可以用作替代碳水化合物原料。该项目将重点关注微生物群落的发展，作为利用这些蔗糖分泌蓝细菌的生产能力的一种手段。 在这种方法中，分泌糖的蓝细菌将与酵母菌株共培养，酵母菌株可以有效地利用碳水化合物并将其生物转化为商品，而无需蔗糖纯化或加工。酵母菌强劲生长和利用蓝藻糖的能力的提高也将最终提高蔗糖向生物燃料和生物产品的转化。 为此，将开发一个模型微生物群落，其中工程酵母将利用光合作用产生的蔗糖来生产脂肪醇、一种有用的表面活性剂和潜在的生物燃料添加剂。 该项目包括通过密歇根州立大学植物基因组学项目和科拉纳学者项目为研究生和本科生提供研究和教育经验。 此外，该项目还涉及参与斯库克拉夫特学院的本科生研究项目，以及通过斯隆学院赞助的德尔菲政策研究，就合成生物学在生物技术应用中的应用进行科学政策讨论。 技术描述该项目的总体目标是开发一种共培养系统，利用蓝细菌和酵母工程菌株，利用二氧化碳和光过量生产蔗糖。该方法依赖于通过共培养进行蔗糖的两步光生物学生产，并将提供关于如何为此目的设计物种间相互作用的基本理解。该项目将利用 PI 实验室最近开发的工程化蓝藻菌株，细长聚球藻 PCC 7942 (S. elongatus)，该菌株基因稳定，能够将多达 85% 的固定碳重新引导至排泄蔗糖。 该菌株具有较高的光合作用生产比活性，因此如果规模化培养能够缓解三个挑战，那么作为替代碳水化合物原料来源具有相当大的潜力，这将通过拟议的研究来解决。 这些挑战是：1）将反应器的复杂性和成本降至与商品产品价值一致的程度，2）降低纯化、加工和将蓝藻碳水化合物输送到下游应用的成本，以及3）降低光生物反应器污染的风险。 为此，该项目将重点开发细长酵母与酿酒酵母（酵母）的直接共培养，从而灵活地将蔗糖转化为附加值化合物。 该项目将探索改善共培养中酵母生长的具体策略，包括改善酵母碳吸收、提高酵母对蓝藻光合代谢副产物的耐受性，以及通过工程化物理方法增强从蓝藻到酵母的碳通量和特异性。协会。将评估这些策略，以提高酵母的稳健性和在共培养中生长时来自 S. elongatus 的碳水化合物的转化效率，并提出通过使用已建立的脂肪醇生产鲸蜡醇的最终原理验证生产酵母菌株。 总体而言，该项目有可能通过为代谢途径工程提供替代方法或通过改进有吸引力的藻类原料来影响藻类生物燃料领域。该项目包括通过密歇根州立大学植物基因组学项目和科拉纳学者项目为研究生和本科生提供研究和教育经验。 此外，该项目还涉及斯隆学院赞助的德尔菲政策研究，参与斯库克拉夫特学院的本科生研究计划，以及关于合成生物学在生物技术应用中的应用的科学政策讨论。