Two-Step Adaptive Laboratory Evolution Enhances Osmotolerance in Engineered Escherichia coli for Improved Succinate Biosynthesis

IF 3.2 3区生物学 Q2 BIOCHEMICAL RESEARCH METHODS

Biotechnology Journal Pub Date : 2025-04-15 DOI:10.1002/biot.70021

Yanzhe Shang, Zhengtong Zhu, Junru Sun, Peng Fei, Yuanchan Luo, Hui Wu

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Abstract

Succinic acid (SA) is a promising platform chemical with broad applications in agricultural, food, and pharmaceutical industries. Microbial production of SA using Escherichia coli typically requires alkaline neutralizers to maintain pH during fermentation, leading to elevated osmotic pressure that severely inhibits SA production. The strain ZZT215, evolved from AFP111 using two-step adaptive laboratory evolution (ALE) strategy, exhibited the improved Na⁺ tolerance and SA productivity without further genetic modification. In 5 L bioreactor fermentation, ZZT215 accumulated 87.02 g/L of SA with a productivity at 1.01 g/(L·h), representing 24.9% and 21.7% increases compared to the parent strain AFP111, respectively. Transcriptomic analysis revealed downregulated TCA cycle genes and upregulated ABC transporters, indicating metabolic adaptation to osmotic stress. These findings highlight the potential of multiple-step ALE for engineering robust microbial cell factories for SA and other high-value chemicals.

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两步适应性实验室进化增强工程大肠杆菌的渗透耐受性，以改善琥珀酸盐的生物合成

丁二酸（SA）是一种前景广阔的平台化学品，在农业、食品和制药行业有着广泛的应用。利用大肠杆菌通过微生物生产琥珀酸通常需要碱性中和剂来维持发酵过程中的 pH 值，从而导致渗透压升高，严重抑制琥珀酸的生产。采用两步适应性实验室进化（ALE）策略从 AFP111 进化而来的菌株 ZZT215 无需进一步的基因修饰就能表现出更好的 Na+ 耐受性和 SA 生产率。在 5 L 生物反应器发酵过程中，ZZT215 积累了 87.02 g/L 的 SA，生产率为 1.01 g/（L-h），与亲本菌株 AFP111 相比分别提高了 24.9% 和 21.7%。转录组分析表明，TCA 循环基因下调，ABC 转运体上调，表明代谢适应了渗透胁迫。这些发现凸显了多步 ALE 在设计生产 SA 和其他高价值化学品的强大微生物细胞工厂方面的潜力。

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来源期刊

Biotechnology Journal Biochemistry, Genetics and Molecular Biology-Molecular Medicine

CiteScore

8.90

自引率

2.10%

发文量

123

审稿时长

1.5 months

期刊介绍： Biotechnology Journal (2019 Journal Citation Reports: 3.543) is fully comprehensive in its scope and publishes strictly peer-reviewed papers covering novel aspects and methods in all areas of biotechnology. Some issues are devoted to a special topic, providing the latest information on the most crucial areas of research and technological advances. In addition to these special issues, the journal welcomes unsolicited submissions for primary research articles, such as Research Articles, Rapid Communications and Biotech Methods. BTJ also welcomes proposals of Review Articles - please send in a brief outline of the article and the senior author''s CV to the editorial office. BTJ promotes a special emphasis on: Systems Biotechnology Synthetic Biology and Metabolic Engineering Nanobiotechnology and Biomaterials Tissue engineering, Regenerative Medicine and Stem cells Gene Editing, Gene therapy and Immunotherapy Omics technologies Industrial Biotechnology, Biopharmaceuticals and Biocatalysis Bioprocess engineering and Downstream processing Plant Biotechnology Biosafety, Biotech Ethics, Science Communication Methods and Advances.