Enhancement of abscisic acid biosynthesis in Saccharomyces cerevisiae via multidimensional engineering

IF 3.7 3区生物学 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY

Process Biochemistry Pub Date : 2024-09-30 DOI:10.1016/j.procbio.2024.09.029

Xiaofei Song , Jianze Zhang , Xikai Wang , Haonan Yu , Nuo Xu , Longyu Cao , Xiuwen Zhong , Puhong Yi , Jie Sun , Kun Wang , Chao Feng , Weixia Wang , Tingheng Zhu

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引用次数: 0

Abstract

Abscisic acid (ABA), a type of sesquiterpenoid plant hormone, has high application value in agriculture, nutrition and medicine. Herein, we constructed an efficient ABA-producing yeast cell factory by combining multidimensional engineering strategies. Starting from a suitable strain, YS010 was selected from 11 varieties of S. cerevisiae strains by evaluating ergosterol content and growth ability, then the biosynthetic pathway of ABA derived from Botris cinerea was constructed, resulting in 1.93 mg L⁻¹ ABA. Next, the metabolic flux of the mevalonic acid (MVA) pathway was increased to enhance the synthesis of the precursor farnesyl pyrophosphate (FPP). To further increase the FPP competitiveness of the ABA synthesis pathway, we attempted to enhance the catalytic performance of BcABA3 through enzyme engineering, and the ABA yield of the mutant strain YS036-ABAP^A206D reached 2.64 mg L⁻¹ in SC-ura medium. In addition, we developed a multi-copy integration strategy, TPI1-delta driven CRISPR-Cas9 (TDI-CRISPR) integration system, to realize the high copy and stable expression of bcaba1, bcaba2 and bcaba3, which enabled the titer of ABA to reach 17.47 mg L⁻¹. Finally, by optimising the fermentation medium, the ABA titer reached 30.30 mg L⁻¹, which was the highest level ever reported for de novo ABA biosynthesis in S. cerevisiae.

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通过多维工程提高赤霉酸在酿酒酵母中的生物合成能力

脱落酸（ABA）是一种倍半萜类植物激素，在农业、营养学和医学方面具有很高的应用价值。在此，我们结合多维工程策略，构建了一个高效的ABA生产酵母细胞工厂。首先，通过评估麦角甾醇含量和生长能力，从 11 个 S. cerevisiae 菌株品种中筛选出一株合适的菌株 YS010，然后构建了从 Botris cinerea 提取 ABA 的生物合成途径，得到 1.93 mg L-1 ABA。接着，增加了甲羟戊酸（MVA）途径的代谢通量，以提高前体焦磷酸法尼基（FPP）的合成。为了进一步提高 FPP 在 ABA 合成途径中的竞争力，我们尝试通过酶工程提高 BcABA3 的催化性能，结果突变株 YS036-ABAPA206D 在 SC-ura 培养基中的 ABA 产量达到了 2.64 mg L-1。此外，我们还开发了一种多拷贝整合策略--TPI1-delta驱动的CRISPR-Cas9（TDI-CRISPR）整合系统，实现了bcaba1、bcaba2和bcaba3的高拷贝和稳定表达，使ABA的滴度达到17.47 mg L-1。最后，通过优化发酵培养基，ABA 的滴度达到了 30.30 mg L-1，这是迄今为止报道的 S. cerevisiae 从头合成 ABA 的最高水平。

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来源期刊

Process Biochemistry 生物-工程：化工

CiteScore

8.30

自引率

4.50%

发文量

374

审稿时长

53 days

期刊介绍： Process Biochemistry is an application-orientated research journal devoted to reporting advances with originality and novelty, in the science and technology of the processes involving bioactive molecules and living organisms. These processes concern the production of useful metabolites or materials, or the removal of toxic compounds using tools and methods of current biology and engineering. Its main areas of interest include novel bioprocesses and enabling technologies (such as nanobiotechnology, tissue engineering, directed evolution, metabolic engineering, systems biology, and synthetic biology) applicable in food (nutraceutical), healthcare (medical, pharmaceutical, cosmetic), energy (biofuels), environmental, and biorefinery industries and their underlying biological and engineering principles.