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Probing the Synergistic Ratio of P450/CPR To Improve (+)-Nootkatone Production in Saccharomyces cerevisiae
Journal of Agricultural and Food Chemistry ( IF 5.7 ) Pub Date : 2022-01-11 , DOI: 10.1021/acs.jafc.1c07035 Yaping Cha 1 , Wen Li 1 , Tao Wu 1 , Xia You 1 , Hefeng Chen 1 , Chaoyi Zhu 1 , Min Zhuo 1 , Bo Chen 2 , Shuang Li 1
Journal of Agricultural and Food Chemistry ( IF 5.7 ) Pub Date : 2022-01-11 , DOI: 10.1021/acs.jafc.1c07035 Yaping Cha 1 , Wen Li 1 , Tao Wu 1 , Xia You 1 , Hefeng Chen 1 , Chaoyi Zhu 1 , Min Zhuo 1 , Bo Chen 2 , Shuang Li 1
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(+)-Nootkatone is an expensive sesquiterpene substance found in grapefruit peels and the heartwood of yellow cedar. It can be used as a food additive, perfume, and insect repellent; therefore, its highly efficient production is greatly needed. However, the low catalytic efficiency of the membrane-anchored cytochrome P450/P450 reductase system (HPO/AtCPR) is the main challenge and limits the production of (+)-nootkatone. We developed an effective high-throughput screening system based on cell wall destruction to probe the optimal ratio of HPO/AtCPR, which achieved a twofold elevation in (+)-valencene oxidation in Saccharomyces cerevisiae. An engineered strain PK2RI-AtC/Hm6A was constructed to realize de novo (+)-nootkatone production by a series of metabolic engineering strategies. In biphasic fed-batch fermentation, maximum titers of 3.73 and 1.02 g/L for (+)-valencene and (+)-nootkatone, respectively, were achieved. The dramatically improved performance of the constructed S. cerevisiae provides an excellent approach for economical production of (+)-nootkatone from glucose.
中文翻译：
探讨 P450/CPR 提高酿酒酵母 (+)-Nootkatone 产量的协同比
(+)-Nootkatone 是一种昂贵的倍半萜烯物质，存在于葡萄柚皮和黄雪松的心材中。可用作食品添加剂、香料、驱虫剂；因此，非常需要它的高效生产。然而，膜锚定细胞色素 P450/P450 还原酶系统 (HPO/AtCPR) 的低催化效率是主要挑战并限制了 (+)-nootkatone 的生产。我们开发了一种基于细胞壁破坏的有效高通量筛选系统来探索 HPO/AtCPR 的最佳比例，该系统在酿酒酵母中实现 (+)-朱栾倍半萜氧化的两倍提升。工程菌株 PK2RI-AtC/H m6A 被构建为通过一系列代谢工程策略实现从头 (+)-nootkatone 生产。在双相补料分批发酵中，(+)-朱栾倍半萜和 (+)-诺卡酮的最大滴度分别达到 3.73 和 1.02 g/L。构建的酿酒酵母显着提高的性能为从葡萄糖中经济地生产 (+)-nootkatone 提供了一种极好的方法。
更新日期：2022-01-26
中文翻译：
探讨 P450/CPR 提高酿酒酵母 (+)-Nootkatone 产量的协同比
(+)-Nootkatone 是一种昂贵的倍半萜烯物质，存在于葡萄柚皮和黄雪松的心材中。可用作食品添加剂、香料、驱虫剂；因此，非常需要它的高效生产。然而，膜锚定细胞色素 P450/P450 还原酶系统 (HPO/AtCPR) 的低催化效率是主要挑战并限制了 (+)-nootkatone 的生产。我们开发了一种基于细胞壁破坏的有效高通量筛选系统来探索 HPO/AtCPR 的最佳比例，该系统在酿酒酵母中实现 (+)-朱栾倍半萜氧化的两倍提升。工程菌株 PK2RI-AtC/H m6A 被构建为通过一系列代谢工程策略实现从头 (+)-nootkatone 生产。在双相补料分批发酵中，(+)-朱栾倍半萜和 (+)-诺卡酮的最大滴度分别达到 3.73 和 1.02 g/L。构建的酿酒酵母显着提高的性能为从葡萄糖中经济地生产 (+)-nootkatone 提供了一种极好的方法。
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