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Efficient Biosynthesis of (2S)‑Eriodictyol from (2S)‑Naringenin in Saccharomyces cerevisiae through a Combination of Promoter Adjustment and Directed Evolution

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posted on 23.11.2020, 20:34 by Song Gao, Xiaoyu Xu, Weizhu Zeng, Sha Xu, Yunbin Lyv, Yue Feng, Guoyin Kai, Jingwen Zhou, Jian Chen
The compound (2S)-eriodictyol is an important flavonoid that can be derived from (2S)-naringenin through flavonoid 3′-hydroxylase (F3′H) catalyzation. F3′H is a cytochrome P450 enzyme that requires a cytochrome P450 reductase (CPR) to function. However, P450s have limited applications in industrial scale biosynthesis, owing to their low activity. Here, an efficient SmF3′H and a matched SmCPR were identified from Silybum marianum. To improve the efficiency of SmF3′H, we established a high-throughput detection method for (2S)-eriodictyol, in which the promoter combination of SmF3′H and SmCPR were optimized in Saccharomyces cerevisiae. The results revealed that SmF3′H/SmCPR should be expressed by using promoters with similar and strong expression levels. Furthermore, directed evolution was applied to further improve the efficiency of SmF3′H/SmCPR. With the optimized promoter and mutated combinations SmF3′HD285N/SmCPRI453V, the (2S)-eriodictyol titer was improved to 3.3 g/L, the highest titer in currently available reports. These results indicated that S. cerevisiae is an ideal platform for functional expression of flavonoid related P450 enzymes.

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