{"title":"三重-CRISPRi-介导的莽草酸途径分支基因下调，促进酿酒酵母中 2-PE 的生物合成","authors":"Shuaihu Fang, Xianghao Fan, Jinlan Li, Zijing Zhu, Pingping Huang, Xiaobao Qi","doi":"10.1007/s00217-023-04461-0","DOIUrl":null,"url":null,"abstract":"<div><p>The 2-phenylethanol biosynthesis in <i>Saccharomyces cerevisiae</i> is limited by multiple-branch metabolism in the shikimate pathway. In this research, a total of 4 × 4 (sites × genes) guide sequences from four branch genes (<i>TYR1</i>, <i>ARO8</i>, <i>AAT2</i> and <i>ALD3</i>) were designed. A single-gene down-regulation library of 4 × 4 <i>Saccharomyces cerevisiae</i> strains was constructed. By the assessment of gene expression level and 2-phenylethanol production, the optimal guide sequences of <i>TYR1</i>/<i>AAT2</i>/<i>ALD3</i> were identified. On these bases, we first developed a high-yielding 2-phenylethanol strain carrying triple-CRISPRi system for simultaneous three branch repression. The INVScI-<i>TYR1</i>.<i>AAT2</i>.<i>ALD3</i> successfully achieved the desired transcriptional repression effect and led to a 1.89-fold increase in 2-phenylethanol production compared to the starting strain. Triple-CRISPRi-mediated down-regulation of the shikimate pathway branch genes provided a convenient and efficient solution for the development of 2-phenylethanol high-yield <i>Saccharomyces cerevisiae</i> engineering strain.</p><h3>Graphical abstract</h3><div><figure><div><div><picture><source><img></source></picture></div></div></figure></div></div>","PeriodicalId":549,"journal":{"name":"European Food Research and Technology","volume":"250 7","pages":"1881 - 1890"},"PeriodicalIF":3.0000,"publicationDate":"2024-05-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Triple-CRISPRi-mediated down-regulation of the shikimate pathway branch genes for enhancing 2-PE biosynthesis in Saccharomyces cerevisiae\",\"authors\":\"Shuaihu Fang, Xianghao Fan, Jinlan Li, Zijing Zhu, Pingping Huang, Xiaobao Qi\",\"doi\":\"10.1007/s00217-023-04461-0\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>The 2-phenylethanol biosynthesis in <i>Saccharomyces cerevisiae</i> is limited by multiple-branch metabolism in the shikimate pathway. In this research, a total of 4 × 4 (sites × genes) guide sequences from four branch genes (<i>TYR1</i>, <i>ARO8</i>, <i>AAT2</i> and <i>ALD3</i>) were designed. A single-gene down-regulation library of 4 × 4 <i>Saccharomyces cerevisiae</i> strains was constructed. By the assessment of gene expression level and 2-phenylethanol production, the optimal guide sequences of <i>TYR1</i>/<i>AAT2</i>/<i>ALD3</i> were identified. On these bases, we first developed a high-yielding 2-phenylethanol strain carrying triple-CRISPRi system for simultaneous three branch repression. The INVScI-<i>TYR1</i>.<i>AAT2</i>.<i>ALD3</i> successfully achieved the desired transcriptional repression effect and led to a 1.89-fold increase in 2-phenylethanol production compared to the starting strain. Triple-CRISPRi-mediated down-regulation of the shikimate pathway branch genes provided a convenient and efficient solution for the development of 2-phenylethanol high-yield <i>Saccharomyces cerevisiae</i> engineering strain.</p><h3>Graphical abstract</h3><div><figure><div><div><picture><source><img></source></picture></div></div></figure></div></div>\",\"PeriodicalId\":549,\"journal\":{\"name\":\"European Food Research and Technology\",\"volume\":\"250 7\",\"pages\":\"1881 - 1890\"},\"PeriodicalIF\":3.0000,\"publicationDate\":\"2024-05-02\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"European Food Research and Technology\",\"FirstCategoryId\":\"97\",\"ListUrlMain\":\"https://link.springer.com/article/10.1007/s00217-023-04461-0\",\"RegionNum\":3,\"RegionCategory\":\"农林科学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"FOOD SCIENCE & TECHNOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"European Food Research and Technology","FirstCategoryId":"97","ListUrlMain":"https://link.springer.com/article/10.1007/s00217-023-04461-0","RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"FOOD SCIENCE & TECHNOLOGY","Score":null,"Total":0}
Triple-CRISPRi-mediated down-regulation of the shikimate pathway branch genes for enhancing 2-PE biosynthesis in Saccharomyces cerevisiae
The 2-phenylethanol biosynthesis in Saccharomyces cerevisiae is limited by multiple-branch metabolism in the shikimate pathway. In this research, a total of 4 × 4 (sites × genes) guide sequences from four branch genes (TYR1, ARO8, AAT2 and ALD3) were designed. A single-gene down-regulation library of 4 × 4 Saccharomyces cerevisiae strains was constructed. By the assessment of gene expression level and 2-phenylethanol production, the optimal guide sequences of TYR1/AAT2/ALD3 were identified. On these bases, we first developed a high-yielding 2-phenylethanol strain carrying triple-CRISPRi system for simultaneous three branch repression. The INVScI-TYR1.AAT2.ALD3 successfully achieved the desired transcriptional repression effect and led to a 1.89-fold increase in 2-phenylethanol production compared to the starting strain. Triple-CRISPRi-mediated down-regulation of the shikimate pathway branch genes provided a convenient and efficient solution for the development of 2-phenylethanol high-yield Saccharomyces cerevisiae engineering strain.
期刊介绍：
The journal European Food Research and Technology publishes state-of-the-art research papers and review articles on fundamental and applied food research. The journal''s mission is the fast publication of high quality papers on front-line research, newest techniques and on developing trends in the following sections:
-chemistry and biochemistry-
technology and molecular biotechnology-
nutritional chemistry and toxicology-
analytical and sensory methodologies-
food physics.
Out of the scope of the journal are:
- contributions which are not of international interest or do not have a substantial impact on food sciences,
- submissions which comprise merely data collections, based on the use of routine analytical or bacteriological methods,
- contributions reporting biological or functional effects without profound chemical and/or physical structure characterization of the compound(s) under research.
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