合成生物学 ›› 2022, Vol. 3 ›› Issue (6): 1174-1200.DOI: 10.12211/2096-8280.2022-022

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合成生物技术驱动酪丁酸梭菌细胞工厂开发的研究进展

刘家宇1, 杨智晗2, 杨蕾2, 朱丽英3, 朱政明1, 江凌1,4   

  1. 1.南京工业大学食品与轻工学院,江苏 南京 211816
    2.南京工业大学生物与制药工程学院,江苏 南京 211816
    3.南京工业大学化学与分子工程学院,江苏 南京 211816
    4.南京工业大学材料化学工程国家重点实验室,江苏 南京 211816
  • 收稿日期:2022-04-14 修回日期:2022-05-25 出版日期:2022-12-31 发布日期:2023-01-17
  • 通讯作者: 朱政明,江凌
  • 作者简介:刘家宇(1998—),男,硕士研究生。研究方向为微生物系统与合成生物学。E-mail:liujoy@njtech.edu.cn
    朱政明(1989—),男,博士,副教授。研究方向为微生物系统与合成生物学。E-mail:zhuzm@njtech.edu.cn
    江凌(1982—),男,博士,教授。研究方向为发酵工程与酶工程。E-mail:jiangling@njtech.edu.cn
  • 基金资助:
    国家自然科学基金优秀青年科学基金项目(31922070);国家自然科学基金青年科学基金项目(22008114);国家自然科学基金山东联合基金重点项目(U2106228);江苏省自然科学基金青年基金项目(BK20200684);江苏省先进生物制造创新中心资助(XTC2205)

Advances in the development of Clostridium tyrobutyricum cell factories driven by synthetic biotechnology

Jiayu LIU1, Zhihan YANG2, Lei YANG2, Liying ZHU3, Zhengming ZHU1, Ling JIANG1,4   

  1. 1.School of Food and Light Industry,Nanjing Tech University,Nanjing 211816,Jiangsu,China
    2.School of Biological and Pharmaceutical Engineering,Nanjing Tech University,Nanjing 211816,Jiangsu,China
    3.School of Chemical and Molecular Engineering,Nanjing Tech University,Nanjing 211816,Jiangsu,China
    4.State Key Laboratory of Chemical Engineering of Materials,Nanjing Tech University,Nanjing 211816,Jiangsu,China
  • Received:2022-04-14 Revised:2022-05-25 Online:2022-12-31 Published:2023-01-17
  • Contact: Zhengming ZHU, Ling JIANG

摘要:

作为一种重要的工业微生物和新型益生菌,酪丁酸梭菌是厌氧条件下代谢多种底物产生丁酸的优势菌株,在其他精细化学品生产和大健康领域亦具有广泛应用。然而,获取生产效率高、鲁棒性强的高版本酪丁酸梭菌细胞工厂,仍然面临着遗传转化效率低、遗传操作工具有限、调控手段单一等诸多挑战。近年来,随着合成生物学的不断发展和酪丁酸梭菌生物信息数据的逐步完善,多种研究策略和技术,包括基因编辑系统等,被用于设计和构筑酪丁酸梭菌底盘细胞高效合成各种精细化学品。本文首先对酪丁酸梭菌独特的生理特性进行了概述。然后,对酪丁酸梭菌底盘细胞改造过程中涉及的系统生物学方法以及遗传操作工具的构建方法与技术进行了总结。同时,探讨了酪丁酸梭菌中多类型代谢调控策略以及群体感应系统的开发及其在合成精细化学品中的应用。最后,从遗传转化效率提升、基因编辑工具拓展、基因回路设计与重构高通量筛选平台建立、一碳气体利用等方面对酪丁酸梭菌底盘细胞的创制进行了展望。

关键词: 酪丁酸梭菌, 底盘细胞, 合成生物技术, 系统生物学, 代谢工程

Abstract:

As an important industrial microorganism and a novel probiotic, Clostridium tyrobutyricum is a superior strain for metabolizing various substrates to produce butyric acid under anaerobic condition, presenting a great potential for the valorization of agricultural wastes. Consequently, this bacterium with high yield of butyric acid has also been widely used in other fields, such as fine chemical production and human health. However, there are still many challenges in the construction of highly productive and robust C. tyrobutyricum cell factories. For example, the genetic transformation efficiency is rather low, due to the presence of restriction-modification systems. Gene editing tools are less developed and strain construction suffers from tedious processes and low efficiency. Moreover, genetic modification of C. tyrobutyricum is limited to a single mode of metabolic regulation, either knockout or overexpression, which is far behind the conventional model hosts. In recent years, with the continuous rapid development of synthetic biology and the collection of increasing amounts of C. tyrobutyricum bioinformatics data, a variety of research strategies and techniques, particularly the gene editing systems, have been employed to design and construct C. tyrobutyricum cell factories for efficient production of various fine chemicals. In this paper, we firstly provide an overview of the unique physiological properties of C. tyrobutyricum, including substrate range, environmental adaptability, butyric acid synthesis pathway, as well as the one-carbon gas fixation and energy metabolism pathways. Subsequently, we summarize the systems biology methods as well as the genetic manipulation tools for the modification of C. tyrobutyricum chassis cell, such as biological elements, the conjugation system, and the CRISPR/Cas system. Meanwhile, we discuss the static and dynamic metabolic regulation strategies and two types of quorum sensing systems (agr-type andRRNPP-type) as well as their applications in the synthesis of fine chemicals in C. tyrobutyricum. Finally, we prospect the trends for the creation of C. tyrobutyricum chassis cell, in terms of enhancing genetic transformation efficiency, expanding gene editing tools, designing and reconstructing gene circuits, establishing high-throughput screening platforms, and utilizing one-carbon gas.

Key words: Clostridium tyrobutyricum, chassis cells, synthetic biotechnology, systems biology, metabolic engineering

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