CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
University of Chinese Academy of Sciences, Beijing 100049, China.
J Agric Food Chem. 2024 Mar 27;72(12):6463-6470. doi: 10.1021/acs.jafc.3c08723. Epub 2024 Mar 19.
Eugenol, the main component of essential oil from the clove tree, has great potential as an alternative bioresource feedstock for biosynthesis purposes. Although eugenol degradation to ferulic acid was investigated, an efficient method for directly converting eugenol to targeted natural products has not been established. Herein we identified the inherent inhibitions by simply combining the previously reported ferulic acid biosynthetic pathway and vanillin biosynthetic pathway. To overcome this, we developed a novel biosynthetic pathway for converting eugenol into vanillin, by introducing cinnamoyl-CoA reductase (CCR), which catalyzes conversion of coniferyl aldehyde to feruloyl-CoA. This approach bypasses the need for two catalysts, namely coniferyl aldehyde dehydrogenase and feruloyl-CoA synthetase, thereby eliminating inhibition while simplifying the pathway. To further improve efficiency, we enhanced CCR catalytic efficiency via directed evolution and leveraged an artificialvanillin biosensor for high-throughput screening. Switching the cofactor preference of CCR from NADP to NAD significantly improved pathway efficiency. This newly designed pathway provides an alternative strategy for efficiently biosynthesizing feruloyl-CoA-derived natural products using eugenol.
丁香酚是丁香树精油的主要成分，具有作为生物合成替代生物资源原料的巨大潜力。尽管已经研究了丁香酚降解为阿魏酸，但尚未建立将丁香酚直接转化为目标天然产物的有效方法。在此，我们通过简单地组合先前报道的阿魏酸生物合成途径和香草醛生物合成途径，鉴定出了固有的抑制作用。为了克服这一问题，我们通过引入肉桂酰辅酶 A 还原酶（CCR），开发了一种将丁香酚转化为香草醛的新型生物合成途径，该酶可催化松柏醛转化为阿魏酰辅酶 A。该方法避免了需要两种催化剂（即松柏醛脱氢酶和阿魏酰辅酶 A 合成酶），从而消除了抑制作用并简化了途径。为了进一步提高效率，我们通过定向进化提高了 CCR 的催化效率，并利用人工香草醛生物传感器进行高通量筛选。将 CCR 的辅因子偏好从 NADP 切换到 NAD 可显著提高途径效率。该新设计的途径为使用丁香酚高效生物合成阿魏酰辅酶 A 衍生的天然产物提供了一种替代策略。
