Expanding the repertoire of aromatic chemicals by microbial production

Microbial production of aromatic chemicals would greatly contribute to solving the problems with fossil resource supply and environmentally sustainable development. Engineering and extending the shikimate/aromatic amino acid biosynthetic pathways are important routes for microbial production of various aromatic chemicals. With advances in metabolic engineering and synthetic biology, we can broaden the product spectrum and obtain several valuable and novel aromatic chemicals from renewable feedstocks. Here, in this review, the latest research progress on microbial production of various aromatic chemicals, and recent metabolic engineering and synthetic biology strategies targeting the central carbon metabolism, the shikimate and aromatic amino acid biosynthetic pathways are summarized and discussed. This work aims to provide some valuable tips for the construction of cost‐effective engineered strains for producing various aromatic chemicals. © 2018 Society of Chemical Industry     

微生物合成2,3-丁二醇的代谢工程

2,3-丁二醇（2,3-BD）是一种重要的微生物代谢产物,广泛应用于食品、医药、化工等多个领域。微生物合成2,3-BD的效率不高一直制约着其生物制造工业化进程,应用代谢工程的理论和方法优化微生物的代谢途径有望解决这一问题。本文全面总结了近年来微生物合成2,3-BD研究过程中的菌株改造和构建技术,包括过表达合成途径中的关键酶编码基因、敲除旁路代谢途径关键酶编码基因、应用辅因子工程手段对天然菌株代谢网络进行重新设计和合理改造,以及利用合成生物学技术在模式菌株中构建全新的代谢途径,实现2,3-BD的高效生物合成。最后,本文对未来的研究方向进行了展望,提出了进一步利用先进的合成生物学方法构建高效细胞工厂的指导性建议。     

High butanol production by regulating carbon,redox and energy in Clostridia? ?

Butanol is a promising biofuel with high energy intensity and can be used as gasoline substitute. It can be produced as a sustainable energy by microorganisms (such as Clostridia) from low-value biomass. However, the low productivity, yield and selectivity in butanol fermentation are still big challenges due to the lack of an efficient butanol-producing host strain. In this article, we systematically review the host cell engineering of Clostridia, focusing on (1) various strategies to rebalance metabolic flux to achieve a high butanol production by regulating the metabolism of carbon, redox or energy, (2) the challenges in pathway manipulation, and (3) the application of proteomics technology to understand the intracellular metabolism. In addition, the process engineering is also briefly described. The objective of this review is to summarize the previous research achievements in the metabolic engineering of Clostridium and provide guidance for future novel strain construction to effectively produce butanol.     

Cell-free synthetic biology in the new era of enzyme engineering

With the gradual rise of enzyme engineering, it has played an essential role in synthetic biology, medicine, and biomanufacturing. However, due to the limitation of the cell membrane, the complexity of cellular metabolism, the difficulty of controlling the reaction environment, and the toxicity of some metabolic products in traditional in vivo enzyme engineering, it is usually problematic to express functional enzymes and produce a high yield of synthesized compounds. Recently, cell-free synthetic biology methods for enzyme engineering have been proposed as alternative strategies. This cell-free method has no limitation of the cell membrane and no need to maintain cell viability, and each biosynthetic pathway is highly flexible. This property makes cell-free approaches suitable for the production of valuable products such as functional enzymes and chemicals that are difficult to synthesize. This article aims to discuss the latest advances in cell-free enzyme engineering, assess the trend of this developing topical filed, and analyze its prospects.     

微生物细胞工厂碳流调控进展

随着代谢工程技术的进步,越来越多微生物细胞工厂可用于化学品发酵生产。微生物细胞生产化学品具有生产条件温和、环境友好等优势,是实现化学品绿色可持续生产的重要手段。为了提高微生物细胞工厂的产量、得率和生产强度,传统代谢工程手段主要采用基因过表达或基因敲除方式增大目标代谢路径碳代谢流。然而由于代谢流调控精度不足,易导致细胞生产能力下降。本文主要针对微生物细胞工厂碳流调控中存在的瓶颈问题,从代谢流改造靶点选择、细胞生长与产物合成碳流平衡、副产物路径与产物合成竞争、产物合成效率强化四个角度,系统综述微生物细胞工厂碳代谢流调控的最新进展。并从高精度、仿生学、智能化、多任务、快响应调控工具的设计出发,对未来微生物细胞工厂的发展趋势进行展望。     

合成生物系统构建与绿色生物“智”造

微生物细胞工厂生产化学品是解决能源和环境问题的有效方式之一。越来越多的化合物可通过合成生物系统实现在微生物中的合成,但菌株的生产能力和鲁棒性仍需进一步提高。提高细胞工厂的智能化,实现生物"智"造过程,将是解决菌株发酵生产能力不足和鲁棒性差的重要途径。本文从蛋白质设计的智能化、生物传感器的智能化、代谢调控的智能化、菌株进化的智能化以及发酵过程智能化等五个层面对生物"智"造的研究现状进行介绍。生物"智"造的发展将为提高工业生物过程的生产水平和过程节能减排做出重要贡献。     

Synthetic biological system construction and green intelligent biological manufacturing

The production of chemicals in microbial cell factories is one of the effective ways to solve energy and environmental problems. Many chemical products have been produced by synthetic biology, but production ability and robustness of the strains still needs to been improved. Development of intelligent cell factory and realizing intelligent biological manufacturing is an important approach to improve the production and robustness. This review introduces the research status of intelligent biological manufacturing from five following aspects: protein design, biosensor, metabolism regulation, strain evolution and fermentation process. The development of biological “intelligence” will make an important contribution to improving the production level of industrial biological processes and process energy saving and emission reduction.     

甲基营养型大肠杆菌构建策略的研究进展

利用微生物细胞工厂实现原料转化和产品合成是绿色生物制造的核心。然而,当前生物制造仍以富含糖类的谷物粮食为主要原料,存在“与民争粮”的争议,亟需开发非粮原料。甲醇作为煤化工产业中的重要产品,具有来源广、价格低、还原性强等优势,有替代粮食原料的潜力。天然甲基营养菌可利用甲醇生产单细胞蛋白和各种氨基酸,但存在理论收率低、遗传改造工具不足等缺点。随着合成生物学的发展,以大肠杆菌等模式生物作为底盘细胞构建人工甲基营养菌,实现甲醇到各种化学品的生产已成为研究热点。本文总结了多种甲基营养型大肠杆菌的构建策略,明确了影响天然路径代谢的关键因素与代谢过程中的重要中间产物,概括了各种天然路径在大肠杆菌中的优化策略与人工新路径的构建方法,并对工程菌株的优化提出了展望。     

Cell-free synthetic biology in the new era of enzyme engineering

With the gradual rise of enzyme engineering, it has played an essential role in synthetic biology, medicine, and biomanufacturing. However, due to the limitation of the cell membrane, the complexity of cellular metabolism, the difficulty of controlling the reaction environment, and the toxicity of some metabolic products in traditional in vivo enzyme engineering, it is usually problematic to express functional enzymes and produce a high yield of synthesized compounds. Recently, cell-free synthetic biology methods for enzyme engineering have been proposed as alternative strategies. This cell-free method has no limitation of the cell membrane and no need to maintain cell viability, and each biosynthetic pathway is highly flexible. This property makes cell-free approaches suitable for the production of valuable products such as functional enzymes and chemicals that are difficult to synthesize. This article aims to discuss the latest advances in cell-free enzyme engineering, assess the trend of this developing topical filed, and analyze its prospects.     

人工合成微生物混菌体系的研究进展

合成生物学正在从设计构建基本功能元件和模块,逐步向着从头设计人工细胞及构建人工生物群落的方向发展,人工合成微生物混菌体系已经成为未来合成生物学研究的重要方向。本文综述了人工构建微生物群落生态关系、群落时空动态和分布式计算等基础研究的进展。同时,微生物混菌体系在医药、环境、能源等领域发挥着不可替代的作用,人工合成混菌体系在相关领域也表现出巨大的应用潜力。     

Engineering Bacterial Cellulose by Synthetic Biology

Synthetic biology is an advanced form of genetic manipulation that applies the principles of modularity and engineering design to reprogram cells by changing their DNA. Over the last decade, synthetic biology has begun to be applied to bacteria that naturally produce biomaterials, in order to boost material production, change material properties and to add new functionalities to the resulting material. Recent work has used synthetic biology to engineer several Komagataeibacter strains; bacteria that naturally secrete large amounts of the versatile and promising material bacterial cellulose (BC). In this review, we summarize how genetic engineering, metabolic engineering and now synthetic biology have been used in Komagataeibacter strains to alter BC, improve its production and begin to add new functionalities into this easy-to-grow material. As well as describing the milestone advances, we also look forward to what will come next from engineering bacterial cellulose by synthetic biology.     

半导体合成生物学的研究进展

半导体合成生物学是研究半导体技术与合成生物学之间协同作用的一门交叉学科。其涉及的活细胞-半导体材料杂合体系具有独特的能量和信号转导机制,不仅维持活细胞的代谢能力,而且保留半导体材料的光电学物理特性,在化工、通讯、计算、能源及医疗等领域具有广阔的应用前景。综述了半导体合成生物学在生物催化、智能生物传感以及新型DNA数据存储领域的最新研究进展,讨论了目前研究面临的技术难题及解决方案,旨在为合成生物学和半导体技术这两个影响化工发展的领域提供有价值的参考。     

The Biochemistry of Phytocannabinoids and Metabolic Engineering of Their Production in Heterologous Systems

The medicinal properties of cannabis and the its legal status in several countries and jurisdictions has spurred the massive growth of the cannabis economy around the globe. The value of cannabis stems from its euphoric activity offered by the unique phytocannabinoid tetrahydrocannabinol (THC). However, this is rapidly expanding beyond THC owing to other non-psychoactive phytocannabinoids with new bioactivities that will contribute to their development into clinically useful drugs. The discovery of the biosynthesis of major phytocannabinoids has allowed the exploration of their heterologous production by synthetic biology, which may lead to the industrial production of rare phytocannabinoids or novel synthetic cannabinoid pharmaceuticals that are not easily offered by cannabis plants. This review summarizes the biosynthesis of major phytocannabinoids in detail, the most recent development of their metabolic engineering in various systems, and the engineering approaches and strategies used to increase the yield.     

Recent advancements in bioreactions of cellular and cell-free systems: A study of bacterial cellulose as a model

Conventional approaches of regulating natural biochemical and biological processes are greatly hampered by the complexity of natural systems. Therefore, current biotechnological research is focused on improving biological systems and processes using advanced technologies such as genetic and metabolic engineering. These technologies, which employ principles of synthetic and systems biology, are greatly motivated by the diversity of living organisms to improve biological processes and allow the manipulation and reprogramming of target bioreactions and cellular systems. This review describes recent developments in cell biology, as well as genetic and metabolic engineering, and their role in enhancing biological processes. In particular, we illustrate recent advancements in genetic and metabolic engineering with respect to the production of bacterial cellulose (BC) using the model systems Gluconacetobacter xylinum and Gluconacetobacter hansenii. Besides, the cell-free enzyme system, representing the latest engineering strategies, has been comprehensively described. The content covered in the current review will lead readers to get an insight into developing novel metabolic pathways and engineering novel strains for enhanced production of BC and other bioproducts formation.     

放线菌聚酮类化合物的合成生物学研究及生物制造

聚酮化合物具有广泛的药用活性和极高的经济价值,但如何高效、经济、绿色、环保地合成聚酮化合物是目前急需解决的问题。随着合成生物学的发展及分子生物学技术的进步,不断有新的技术和策略被用于聚酮化合物的生物制造。本文介绍了聚酮化合物生物制造中的关键酶、前体物质及代谢途径等,分析了通过CRISPR技术及翻译后修饰代谢工程优化代谢调控网络;通过替换及优化启动子等手段改造与优化代谢途径;通过构建简单、高效的异源表达系统等策略提高聚酮化合物的生物制造效率等。在此基础上对红霉素、阿维菌素、多杀菌素的合成生物学研究的最新进展进行了总结,进而对当前聚酮化合物生物制造面临的产量及效率低下等问题和可能的解决途径,如平衡初级代谢与次级代谢,构建新型、优势底盘细胞及代谢网络的重新设计与改造等进行了展望。     

Synthetic Multienzyme Assemblies for Natural Product Biosynthesis

In nature, enzymes that catalyze sequential reactions are often assembled as clusters or complexes. The formation of multienzyme complexes, or metabolons, brings the enzyme active sites into proximity to promote intermediate transfer, decrease intermediate leakage, and streamline the metabolic flux towards the desired products. We and others have developed synthetic versions of metabolons through various strategies to enhance the catalytic rates for synthesizing valuable chemicals inside microbes. Synthetic multienzyme complexes range from static enzyme nanostructures to dynamic enzyme coacervates. Enzyme complexation optimizes the metabolic fluxes inside microbes, increases the product titer, and supplies the field with high-yield microbe strains that are amenable to large-scale fermentation. Enzyme complexes constructed inside microbial cells can be separated as independent entities and catalyze biosynthetic reactions ex vivo; such a feature gains these complexes another name, “synthetic organelles” – new subcellular entities with independent structures and functions. Still, the field is seeking new strategies to better balance dynamicity and confinement and to achieve finer control of local compartmentalization in the cells, as the natural multienzyme complexes do. Industrial applications of synthetic multienzyme complexes for the large-scale production of valuable chemicals are yet to be realized. This review focuses on synthetic multienzyme complexes that are constructed and function inside microbial cells.     

他克莫司微生物合成的研究进展

介绍了他克莫司的理化特征和免疫抑制活性,重点阐述了他克莫司发酵法生产的研究现状,着重介绍了他克莫司的合成基因簇及代谢途径研究进展,对他克莫司代谢工程改造进行综述并对今后的研究趋势进行了展望--结合系统生物学为已有的他克莫司菌株进一步进行代谢工程改造提供指导;通过合成生物学构建他克莫司合成途径全新高效的前体关键酶。     

人参皂苷的定向生物转化研究进展

稀有人参皂苷是人参的重要活性成分,但其含量极低或需经过肠道转化才能产生,工业生产中常通过去糖基、脱水等方法将常见人参皂苷转化为稀有人参皂苷。基于此,本文概述了人参皂苷的构效关系及常用转化方法,总结了微生物、酶法定向转化人参皂苷的最新进展,着重介绍了益生菌、食药用真菌转化人参皂苷制备功能食品,以及糖苷酶筛选和组合在提高人参总皂苷转化效率及产率等方面的研究进展;同时探讨了基因工程、溶剂工程、固定化酶等技术对人参皂苷转化效率、产率的影响,展望了蛋白质工程、合成生物学等方法在人参皂苷转化及合成方面的潜在应用价值,为稀有人参皂苷的规模化生产提供了基础。     

合成气的生物利用与定向转化

合成气是来源于石化、煤化工以及生物质加工行业的一类重要原料气体。现有的化学催化路线可将合成气转化为氨、烯烃、甲醇等大宗化工产品,但尚无法实现选择性地合成具有较高附加值的长碳链化合物,而发展合成气的生物转化路线是克服上述难题、拓展产业链的有效策略。本文综述了随着分子遗传操作工具以及合成生物学的快速发展,合成气生物利用相关的菌株代谢工程设计、改造以及发酵工艺优化等方面的研究进展和产业化进程,并指出目前该技术路线在固碳效率、产物合成种类及产量方面还存在不足,亟待优化以满足大规模工业化推广应用的要求。本文还对合成气生物利用与转化的研究现状进行了梳理和总结,并探讨了未来的发展方向,以期为建立具有经济竞争力的合成气生物利用技术和工艺提供参考。     

Construction of microbial cell factories for industrial bioprocesses

The development of industrial microbial processes is gaining unprecedented momentum. Increased concern for environmental issues and the prospect of declining petroleum resources has shifted the industrial focus increasingly to microorganisms as biocatalysts. At the same time systems biology and synthetic biology supply industry and academia with new tools to design optimal microbial cell factories. Among the tools are systems biology approaches allowing the modelling of cellular networks for rational strain design, single cell analyses methods for gaining insight into population hetereogeneity, and an exciting combination of tools from structural biology and synthetic biology, permitting the catalysis of new (unnatural) enzymatic reactions or the production of new (unnatural) chemicals. This perspective article outlines recent advances and new developments within the field of microbial cell factory design. Copyright © 2012 Society of Chemical Industry