PDF(Pubmed)
Abstract：
Resveratrol, a phenylpropanoid compound, exhibits diverse pharmacological properties, making it a valuable candidate for health and disease management. However, the demand for resveratrol exceeds the capacity of plant extraction methods, necessitating alternative production strategies. Microbial synthesis offers several advantages over plant-based approaches and presents a promising alternative. Yarrowia lipolytica stands out among microbial hosts due to its safe nature, abundant acetyl-CoA and malonyl-CoA availability, and robust pentose phosphate pathway. This study aimed to engineer Y. lipolytica for resveratrol production. The resveratrol biosynthetic pathway was integrated into Y. lipolytica by adding genes encoding tyrosine ammonia lyase from Rhodotorula glutinis, 4-coumarate CoA ligase from Nicotiana tabacum, and stilbene synthase from Vitis vinifera. This resulted in the production of 14.3 mg/L resveratrol. A combination of endogenous and exogenous malonyl-CoA biosynthetic modules was introduced to enhance malonyl-CoA availability. This included genes encoding acetyl-CoA carboxylase 2 from Arabidopsis thaliana, malonyl-CoA synthase, and a malonate transporter protein from Bradyrhizobium diazoefficiens. These strategies increased resveratrol production to 51.8 mg/L. The further optimization of fermentation conditions and the utilization of sucrose as an effective carbon source in YP media enhanced the resveratrol concentration to 141 mg/L in flask fermentation. By combining these strategies, we achieved a titer of 400 mg/L resveratrol in a controlled fed-batch bioreactor. These findings demonstrate the efficacy of Y. lipolytica as a platform for the de novo production of resveratrol and highlight the importance of metabolic engineering, enhancing malonyl-CoA availability, and media optimization for improved resveratrol production.
摘要：
白藜芦醇,苯丙素类化合物，表现出不同的药理特性，使其成为健康和疾病管理的宝贵候选人。然而,对白藜芦醇的需求超过了植物提取方法的能力，需要替代生产战略。与基于植物的方法相比，微生物合成具有若干优势，并提出了有希望的替代方案。Yarrowialipolytica由于其安全的性质而在微生物宿主中脱颖而出，丰富的乙酰辅酶A和丙二酰辅酶A可用性，和强大的磷酸戊糖途径。本研究旨在设计Y.Lipolytica用于白藜芦醇生产。白藜芦醇生物合成途径被整合到Y中。通过添加编码来自谷红酵母的酪氨酸氨裂解酶的基因，来自烟草的4-香豆酸CoA连接酶，和葡萄的二苯乙烯合成酶。这导致产生14.3mg/L白藜芦醇。引入内源性和外源性丙二酰辅酶A生物合成模块的组合以增强丙二酰辅酶A的可用性。这包括编码来自拟南芥的乙酰辅酶A羧化酶2的基因，丙二酰辅酶A合成酶，和一种来自重氮根瘤菌的丙二酸转运蛋白。这些策略将白藜芦醇的产量增加到51.8mg/L。发酵条件的进一步优化和利用蔗糖作为YP培养基中的有效碳源，使白藜芦醇在烧瓶发酵中的浓度提高到141mg/L。通过结合这些策略，我们在受控补料分批生物反应器中实现了400mg/L白藜芦醇的滴度。这些发现证明了Y.lipolytica作为从头生产白藜芦醇的平台的功效，并强调了代谢工程的重要性。提高丙二酰辅酶A的可用性，和培养基优化以提高白藜芦醇的产量。
