State Key Laboratory of Subtropical Silviculture, Zhejiang A&F University, Hangzhou 311300, China.
Zhejiang Provincial Key Laboratory of Resources Protection and Innovation of Traditional Chinese Medicine, Zhejiang A&F University, Lin'an, Hangzhou 311300, China.
J Agric Food Chem. 2024 Feb 14;72(6):3077-3087. doi: 10.1021/acs.jafc.3c08529. Epub 2024 Feb 1.
, a microorganism classified as generally recognized as safe for use in the industrial production of food raw materials and additives, has encountered challenges in achieving widespread adoption and popularization as microbial cell factories. These obstacles arise from the intricate nature of manipulating metabolic flux through conventional methods, such as gene knockout and enzyme overexpression. To address this challenge, we developed a CRISPR/dCpf1-based bifunctional regulation system to bidirectionally regulate the expression of multiple genes in . Specifically, through fusing various transcription factors to the C-terminus of dCpf1, the resulting dCpf1-SoxS exhibited both CRISPR interference (CRISPRi) and CRISPR activation (CRISPRa) capabilities in by altering the binding sites of crRNAs. The bifunctional regulation system was used to fine-tune metabolic flux from shikimic acid (SA) and l-serine biosynthesis, resulting in 27-fold and 10-fold increases in SA and l-serine production, respectively, compared to the original strain. These findings highlight the potential of the CRISPR/dCpf1-based bifunctional regulation system in effectively enhancing the yield of target products in .
作为一种被归类为普遍认为安全可用于食品原料和添加剂的工业生产的微生物，已在将其作为微生物细胞工厂广泛采用和推广上面临挑战。这些障碍源于通过传统方法（如基因敲除和酶过表达）操纵代谢通量的复杂性。为了解决这一挑战，我们开发了一种基于 CRISPR/dCpf1 的双功能调控系统，用于双向调控 中的多个基因表达。具体来说，通过将各种转录因子融合到 dCpf1 的 C 末端，所得的 dCpf1-SoxS 通过改变 crRNA 的结合位点，在 中表现出 CRISPR 干扰 (CRISPRi) 和 CRISPR 激活 (CRISPRa) 能力。该双功能调控系统用于微调从莽草酸 (SA) 和 l-丝氨酸生物合成的代谢通量，与原始菌株相比，SA 和 l-丝氨酸的产量分别提高了 27 倍和 10 倍。这些发现突出了基于 CRISPR/dCpf1 的双功能调控系统在有效提高目标产物产量方面的潜力。
