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首页> 外文期刊>Bioprocess and Biosystems Engineering >Enhancing the production of tacrolimus by engineering target genes identified in important primary and secondary metabolic pathways and feeding exogenous precursors
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Enhancing the production of tacrolimus by engineering target genes identified in important primary and secondary metabolic pathways and feeding exogenous precursors

机译：通过在重要的初级和二次代谢途径和喂养外源前体中鉴定的工程靶基因增强凝胶蛋白的产生

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摘要

Tacrolimus has been widely used as a powerful novel immunosuppressant. The objective of this study was to improve the production of tacrolimus by engineering the target genes of important primary and secondary metabolic pathways and feeding exogenous precursors. Based on the metabonomics analysis, the shikimic acid pathway is an important primary metabolic pathway for the producing tacrolimus. Combined overexpression of shikimate kinase and dehydroquinic acid synthetase genes led to a 33.1% enhancement of tacrolimus production compared to parent strain. To predict the most efficient targets in secondary metabolic pathways for improving the production of tacrolimus, a genome-scale dynamic metabolic network model was used. A knockout of the d-lactate dehydrogenase gene, combined with the overexpression of tryptophane synthase and aspartate 1-decarboxylase genes, led to a 29.8% enhancement of tacrolimus production compared to the parent strain. Finally, we investigated the impact of the genetic manipulations on transcription levels, cell growth, cell morphology and production of tacrolimus by qRT-PCR and scanning electron microscopy to reveal the relationship between the growth of strains, the effects of engineering and fermentation. As the efficient synthesis of tacrolimus requires a rich supply of external substrates, the efficiency of the metabolic pathways that convert these substances is extremely important. The combined addition of three external substrates such as shikimic acid, alanine and the n-dodecane increased tacrolimus production by 49.5%. The insights obtained in this study will help further elucidate the mechanisms by which the identified target genes promote the activity of important primary and secondary metabolic pathways for tacrolimus biosynthesis and provide a new feeding strategy to improve tacrolimus production.
机译：他克莫司已被广泛用作强大的新型免疫抑制剂。本研究的目的是通过工程重要初级和次生代谢途径的靶基因来改善躯体司司的生产和喂养外源前体。基于代谢族分析，Shikimic酸途径是生产巨杆菌的重要原始代谢途径。与亲本菌株相比，Shikimate激酶和脱氢喹啉酸合成酶基因的组合过表达导致巨粒蛋白的生产增强了33.1％。为了预测用于改善Tacolimus的产生的二次代谢途径中最有效的靶标，使用了一种基因组级动态代谢网络模型。与色氨酸合酶和天冬氨酸1-脱羧酶基因的过表达结合的D-乳酸脱氢酶基因的敲除，与亲本菌株相比，Tacromus生产的增强的增强的29.8％。最后，通过QRT-PCR和扫描电子显微镜调查了遗传操纵对转录水平，细胞生长，细胞形态和生产的Tacrolimus的影响，揭示了菌株的生长，工程和发酵的影响之间的关系。由于高曲粒的有效合成需要丰富的外部基材供应，转化这些物质的代谢途径的效率非常重要。三种外部基材的组合加入，例如Shikimic酸，丙氨酸和N-十二烷增加了标准蛋白的产量49.5％。本研究中获得的见解将有助于进一步阐明所识别的靶基因促进统计的躯体生物合成的重要原发性和次生代谢途径的机制，并提供新的饲养策略，以改善Tacrolimus生产。

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  • 来源
    《Bioprocess and Biosystems Engineering》 |2019年第7期|1081-1098|共18页
  • 作者

    Li Yang; Liang Shaoxiong; Wang Junhua; Ma Dongxu; Wen Jianping;

  • 作者单位

    Tianjin Univ Minist Educ Key Lab Syst Bioengn Tianjin 300072 Peoples R China|Collaborat Innovat Ctr Chem Sci & Engn Tianjin SynBio Res Platform Tianjin 300072 Peoples R China;

    Huaqiao Univ Coll Lab Chem Engn Xiamen 361021 Fujian Peoples R China;

    Tianjin Univ Minist Educ Key Lab Syst Bioengn Tianjin 300072 Peoples R China|Collaborat Innovat Ctr Chem Sci & Engn Tianjin SynBio Res Platform Tianjin 300072 Peoples R China;

    Tianjin Univ Minist Educ Key Lab Syst Bioengn Tianjin 300072 Peoples R China|Collaborat Innovat Ctr Chem Sci & Engn Tianjin SynBio Res Platform Tianjin 300072 Peoples R China;

    Tianjin Univ Minist Educ Key Lab Syst Bioengn Tianjin 300072 Peoples R China|Collaborat Innovat Ctr Chem Sci & Engn Tianjin SynBio Res Platform Tianjin 300072 Peoples R China;

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  • 正文语种 eng
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  • 关键词

    Tacrolimus; Primary and secondary metabolic pathways; Target genes; qRT-PCR; Scanning electron microscopy; Exogenous substances;

    机译：他克莫司;初级和次生代谢途径;靶基因;QRT-PCR;扫描电子显微镜;外源性物质;

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